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Utilization of a Diazene Core to Target Sulfinic Acid
Title
Utilization of a Diazene Core to Target Sulfinic Acid.
Creator
Morgan, Lexi, Department of Biochemistry
Abstract/Description
Hydrogen peroxide (H2O2) regulates an array of physiological functions and acts as a second messenger at low levels. However, the elevation of these levels can lead to oxidative stress, a state that is implicated in the progression of human diseases, such as cancer and Parkinson's disease. Cysteine is highly susceptible to oxidation from hydrogen peroxide because of the nucleophilicity of the thiol group (R-SH) to form sulfenic acid (RSOH). The thiol group can be further oxidized to form...
Show moreHydrogen peroxide (H2O2) regulates an array of physiological functions and acts as a second messenger at low levels. However, the elevation of these levels can lead to oxidative stress, a state that is implicated in the progression of human diseases, such as cancer and Parkinson's disease. Cysteine is highly susceptible to oxidation from hydrogen peroxide because of the nucleophilicity of the thiol group (R-SH) to form sulfenic acid (RSOH). The thiol group can be further oxidized to form sulfinic acid (RSO2H) and sulfonic acid (RSO3H). Each of these species exhibits unique chemical properties as well as a versatile mechanism to alter protein function. While the regulatory function of sulfenic has been widely studied, very little is known about the role sulfinic acid plays. Mounting evidence suggests that the cysteine sulfinic acid is more regulated than once thought. An enzyme called sulfiredoxin was found to reduce the sulfinic form of certain peroxiredoxins. The discovery of a sulfinic acid reductase suggests a more fundamental role for this modification, thus the proposal of the "sulfinic acid switch" in regard to the protein regulation by hydrogen peroxide. At The Scripps Research Institute, Dr. Kate Carroll's goal is to monitor the oxidation of cysteine through the utilization of novel probes. This project focused on the utilization of a diazene core (R-N=N-R) with a variety of functional groups in order to target sulfinic acid. This electrophilic nitrogen-containing species acts as a specific reagent to target the nucleophilic sulfinic acid. This selective ligation reaction with sulfinic acids has potential utility for detections of oxidative modifications, as well as regulations, in biological systems.
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Date Issued
2013
Identifier
FSU_migr_undergradresearch-0010
Format
Citation
δ/ω-Plectoxin-Pt1a
Title
δ/ω-Plectoxin-Pt1a: An Excitatory Spider Toxin with Actions on both Ca(2+) and Na(+) Channels.
Creator
Zhou, Yi, Zhao, Mingli, Fields, Gregg B., Wu, Chun-Fang, Branton, W.
Abstract/Description
The venom of spider Plectreurys tristis contains a variety of peptide toxins that selectively target neuronal ion channels. O-palmitoylation of a threonine or serine residue, along with a characteristic and highly constrained disulfide bond structure, are hallmarks of a family of toxins found in this venom. Here, we report the isolation and characterization of a new toxin, δ/ω-plectoxin-Pt1a, from this spider venom. It is a 40 amino acid peptide containing an O-palmitoylated Ser-39. Analysis...
Show moreThe venom of spider Plectreurys tristis contains a variety of peptide toxins that selectively target neuronal ion channels. O-palmitoylation of a threonine or serine residue, along with a characteristic and highly constrained disulfide bond structure, are hallmarks of a family of toxins found in this venom. Here, we report the isolation and characterization of a new toxin, δ/ω-plectoxin-Pt1a, from this spider venom. It is a 40 amino acid peptide containing an O-palmitoylated Ser-39. Analysis of δ/ω-plectoxin-Pt1a cDNA reveals a small precursor containing a secretion signal sequence, a 14 amino acid N-terminal propeptide, and a C-terminal amidation signal. The biological activity of δ/ω-plectoxin-Pt1a is also unique. It preferentially blocks a subset of Ca(2+) channels that is apparently not required for neurotransmitter release; decreases threshold for Na(+) channel activation; and slows Na(+) channel inactivation. As δ/ω-plectoxin-Pt1a enhances synaptic transmission by prolonging presynaptic release of neurotransmitter, its effects on Na(+) and Ca(2+) channels may act synergistically to sustain the terminal excitability.
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Date Issued
2013
Identifier
FSU_migr_biomed_faculty_publications-0044
Format
Citation
Personality, Metabolic Rate and Aerobic Capacity
Title
Personality, Metabolic Rate and Aerobic Capacity.
Creator
Terracciano, Antonio, Schrack, Jennifer, Sutin, Angelina, Chan, Wayne, Simonsick, Eleanor M., Ferrucci, Luigi
Abstract/Description
Personality traits and cardiorespiratory fitness in older adults are reliable predictors of health and longevity. We examined the association between personality traits and energy expenditure at rest (basal metabolic rate) and during normal and maximal sustained walking. Personality traits and oxygen (VO(2)) consumption were assessed in 642 participants from the Baltimore Longitudinal Study of Aging. Results indicate that personality traits were mostly unrelated to resting metabolic rate and...
Show morePersonality traits and cardiorespiratory fitness in older adults are reliable predictors of health and longevity. We examined the association between personality traits and energy expenditure at rest (basal metabolic rate) and during normal and maximal sustained walking. Personality traits and oxygen (VO(2)) consumption were assessed in 642 participants from the Baltimore Longitudinal Study of Aging. Results indicate that personality traits were mostly unrelated to resting metabolic rate and energy expenditure at normal walking pace. However, those who scored lower on neuroticism (r = -0.12) and higher on extraversion (r = 0.11), openness (r = 0.13), and conscientiousness (r = 0.09) had significantly higher energy expenditure at peak walking pace. In addition to greater aerobic capacity, individuals with a more resilient personality profile walked faster and were more efficient in that they required less energy per meter walked. The associations between personality and energy expenditure were not moderated by age or sex, but were in part explained by the proportion of fat mass. In conclusion, differences in personality may matter the most during more challenging activities that require cardiorespiratory fitness. These findings suggest potential pathways that link personality to health outcomes, such as obesity and longevity.
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Date Issued
2013
Identifier
FSU_migr_mhs-0011, 10.1371/journal.pone.0054746
Format
Citation
Depressive Symptoms Are Associated with Weight Gain Among Women
Title
Depressive Symptoms Are Associated with Weight Gain Among Women.
Creator
Sutin, Angelina, Zonderman, Alan
Abstract/Description
BACKGROUND: Many studies have linked depression and obesity; few have more than two assessments of depressive symptoms and adiposity to address the potential bidirectional relationship between adiposity and depressive symptoms from young adulthood through old age. We tested whether baseline depressive symptoms are associated with changes in weight, whether baseline adiposity is associated with changes in depressive symptoms, and whether these associations vary by sex.MethodParticipants (n...
Show moreBACKGROUND: Many studies have linked depression and obesity; few have more than two assessments of depressive symptoms and adiposity to address the potential bidirectional relationship between adiposity and depressive symptoms from young adulthood through old age. We tested whether baseline depressive symptoms are associated with changes in weight, whether baseline adiposity is associated with changes in depressive symptoms, and whether these associations vary by sex.MethodParticipants (n=2251; 47% female) were from the Baltimore Longitudinal Study of Aging (BLSA). Using hierarchical linear modeling (HLM) on 30 years of data, the trajectory of adiposity and depressive symptoms over adulthood was estimated from >10 000 observations (mean=4.5 assessments per participant) of body mass index (BMI; kg/m2), waist circumference and hip circumference and >10 000 observations (mean=4.5 assessments per participant) of the Center for Epidemiological Studies Depression Scale (CES-D). Baseline depressive symptoms and adiposity were then tested as predictors of the trajectory of adiposity and depressive symptoms respectively. Additional analyses tested for sex-specific associations. RESULTS: Sex moderated the association between depressive symptoms and weight gain such that women who experienced depressed affect had greater increases in BMI (binteraction=0.12, s.e.=0.04), waist (binteraction=0.22, s.e.=0.10) and hip circumference (binteraction=0.20, s.e.=0.07) across the adult lifespan, controlling for relevant demographic and behavioral covariates. Baseline adiposity was unrelated to the trajectory of depressive symptoms (median b=0.00) for both sexes. CONCLUSIONS: Women who experience symptoms of depression tend to gain more weight across adulthood than men who experience such symptoms. Whether an individual was normal weight or overweight was unrelated to changes in depressive symptoms across adulthood.
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Date Issued
2012
Identifier
FSU_migr_mhs-0012, 10.1017/S0033291712000566
Format
Citation
Kallikrein 6 is a Novel Molecular Trigger of Reactive Astrogliosis
Title
Kallikrein 6 is a Novel Molecular Trigger of Reactive Astrogliosis.
Creator
Scarisbrick, Isobel, Radulovic, Maja, Burda, Joshua, Larson, Nadya, Blaber, Sachiko, Giannini, Caterina, Blaber, Michael, Vandell, Alexander
Abstract/Description
Kallikrein-related peptidase 6 (KLK6) is a trypsin-like serine protease upregulated at sites of central nervous system (CNS) injury, including de novo expression by reactive astrocytes, yet its physiological actions are largely undefined. Taken with recent evidence that KLK6 activates G-protein-coupled protease-activated receptors (PARs), we hypothesized that injury-induced elevations in KLK6 contribute to the development of astrogliosis and that this occurs in a PAR-dependent fashion. Using...
Show moreKallikrein-related peptidase 6 (KLK6) is a trypsin-like serine protease upregulated at sites of central nervous system (CNS) injury, including de novo expression by reactive astrocytes, yet its physiological actions are largely undefined. Taken with recent evidence that KLK6 activates G-protein-coupled protease-activated receptors (PARs), we hypothesized that injury-induced elevations in KLK6 contribute to the development of astrogliosis and that this occurs in a PAR-dependent fashion. Using primary murine astrocytes and the Neu7 astrocyte cell line, we show that KLK6 causes astrocytes to transform from an epitheliod to a stellate morphology and to secrete interleukin 6 (IL-6). By contrast, KLK6 reduced expression of glial fibrillary acidic protein (GFAP). The stellation-promoting activities of KLK6 were shown to be dependent on activation of the thrombin receptor, PAR1, as a PAR1-specific inhibitor, SCH79797, blocked KLK6-induced morphological changes. The ability of KLK6 to promote astrocyte stellation was also shown to be linked to activation of protein kinase C (PKC). These studies indicate that KLK6 is positioned to serve as a molecular trigger of select physiological processes involved in the development of astrogliosis and that this is likely to occur at least in part by activation of the G-protein-coupled receptor, PAR1.
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Date Issued
2012
Identifier
FSU_migr_biomed_faculty_publications-0030, 10.1515/hsz-2011-0241
Format
Citation
Pharmacokinetic Properties of 2(nd)-Generation Fibroblast Growth Factor-1 Mutants for Therapeutic Application
Title
Pharmacokinetic Properties of 2(nd)-Generation Fibroblast Growth Factor-1 Mutants for Therapeutic Application.
Creator
Xia, Xue, Babcock, Joseph, Blaber, Sachiko, Harper, Kathleen, Blaber, Michael
Abstract/Description
Fibroblast growth factor-1 (FGF-1) is an angiogenic factor with therapeutic potential for the treatment of ischemic disease. FGF-1 has low intrinsic thermostability and is characteristically formulated with heparin as a stabilizing agent. Heparin, however, adds a number of undesirable properties that negatively impact safety and cost. Mutations that increase the thermostability of FGF-1 may obviate the need for heparin in formulation and may prove to be useful "2nd-generation" forms for...
Show moreFibroblast growth factor-1 (FGF-1) is an angiogenic factor with therapeutic potential for the treatment of ischemic disease. FGF-1 has low intrinsic thermostability and is characteristically formulated with heparin as a stabilizing agent. Heparin, however, adds a number of undesirable properties that negatively impact safety and cost. Mutations that increase the thermostability of FGF-1 may obviate the need for heparin in formulation and may prove to be useful "2nd-generation" forms for therapeutic use. We report a pharmacokinetic (PK) study in rabbits of human FGF-1 in the presence and absence of heparin, as well as three mutant forms having differential effects upon thermostability, buried reactive thiols, and heparin affinity. The results support the hypothesis that heparan sulfate proteoglycan (HSPG) in the vasculature of liver, kidney and spleen serves as the principle peripheral compartment in the distribution kinetics. The addition of heparin to FGF-1 is shown to increase endocrine-like properties of distribution. Mutant forms of FGF-1 that enhance thermostability or eliminate buried reactive thiols demonstrate a shorter distribution half-life, a longer elimination half-life, and a longer mean residence time (MRT) in comparison to wild-type FGF-1. The results show how such mutations can produce useful 2nd-generation forms with tailored PK profiles for specific therapeutic application.
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Date Issued
2012
Identifier
FSU_migr_biomed_faculty_publications-0041, 10.1371/journal.pone.0048210
Format
Citation
Experimental Support for the Evolution of Symmetric Protein Architecture from a Simple Peptide Motif
Title
Experimental Support for the Evolution of Symmetric Protein Architecture from a Simple Peptide Motif.
Creator
Lee, Jihun, Blaber, Michael
Abstract/Description
The majority of protein architectures exhibit elements of structural symmetry, and "gene duplication and fusion" is the evolutionary mechanism generally hypothesized to be responsible for their emergence from simple peptide motifs. Despite the central importance of the gene duplication and fusion hypothesis, experimental support for a plausible evolutionary pathway for a specific protein architecture has yet to be effectively demonstrated. To address this question, a unique "top-down...
Show moreThe majority of protein architectures exhibit elements of structural symmetry, and "gene duplication and fusion" is the evolutionary mechanism generally hypothesized to be responsible for their emergence from simple peptide motifs. Despite the central importance of the gene duplication and fusion hypothesis, experimental support for a plausible evolutionary pathway for a specific protein architecture has yet to be effectively demonstrated. To address this question, a unique "top-down symmetric deconstruction" strategy was utilized to successfully identify a simple peptide motif capable of recapitulating, via gene duplication and fusion processes, a symmetric protein architecture (the threefold symmetric β-trefoil fold). The folding properties of intermediary forms in this deconstruction agree precisely with a previously proposed "conserved architecture" model for symmetric protein evolution. Furthermore, a route through foldable sequence-space between the simple peptide motif and extant protein fold is demonstrated. These results provide compelling experimental support for a plausible evolutionary pathway of symmetric protein architecture via gene duplication and fusion processes.
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Date Issued
2011
Identifier
FSU_migr_biomed_faculty_publications-0020, 10.1073/pnas.1015032108, PMC3017207
Format
Citation
Human Coronary Artery Smooth Muscle Cell Responses to Bioactive Polyelectrolyte Multilayer Interfaces
Title
Human Coronary Artery Smooth Muscle Cell Responses to Bioactive Polyelectrolyte Multilayer Interfaces.
Creator
Newcomer, Robert, Moussallem, Maroun, Keller, Thomas C. S., Schlenoff, Joseph B., Sang, Qing-Xiang
Abstract/Description
Under normal physiological conditions, mature human coronary artery smooth muscle cells (hCASMCs) exhibit a "contractile" phenotype marked by low rates of proliferation and protein synthesis, but these cells possess the remarkable ability to dedifferentiate into a "synthetic" phenotype when stimulated by conditions of pathologic stress. A variety of polyelectrolyte multilayer (PEMU) films are shown here to exhibit bioactive properties that induce distinct responses from cultured hCASMCs....
Show moreUnder normal physiological conditions, mature human coronary artery smooth muscle cells (hCASMCs) exhibit a "contractile" phenotype marked by low rates of proliferation and protein synthesis, but these cells possess the remarkable ability to dedifferentiate into a "synthetic" phenotype when stimulated by conditions of pathologic stress. A variety of polyelectrolyte multilayer (PEMU) films are shown here to exhibit bioactive properties that induce distinct responses from cultured hCASMCs. Surfaces terminated with Nafion or poly(styrenesulfonic acid) (PSS) induce changes in the expression and organization of intracellular proteins, while a hydrophilic, zwitterionic copolymer of acrylic acid and 3-[2-(acrylamido)-ethyl dimethylammonio] propane sulfonate (PAA-co-PAEDAPS) is resistant to cell attachment and suppresses the formation of key cytoskeletal components. Differential expression of heat shock protein 90 and actin is observed, in terms of both their magnitude and cellular localization, and distinct cytoplasmic patterns of vimentin are seen. The ionophore A23187 induces contraction in confluent hCASMC cultures on Nafion-terminated surfaces. These results demonstrate that PEMU coatings exert direct effects on the cytoskeletal organization of attaching hCASMCs, impeding growth in some cases, inducing changes consistent with phenotypic modulation in others, and suggesting potential utility for PEMU surfaces as a coating for coronary artery stents and other implantable medical devices.
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Date Issued
2011
Identifier
FSU_migr_chm_faculty_publications-0011, 10.4061/2011/854068
Format
Citation
Personality and Metabolic Syndrome
Title
Personality and Metabolic Syndrome.
Creator
Sutin, Angelina, Costa, Paul, Uda, Manuela, Ferrucci, Luigi, Schlessinger, David, Terracciano, Antonio
Abstract/Description
The prevalence of metabolic syndrome has paralleled the sharp increase in obesity. Given its tremendous physical, emotional, and financial burden, it is of critical importance to identify who is most at risk and the potential points of intervention. Psychological traits, in addition to physiological and social risk factors, may contribute to metabolic syndrome. The objective of the present research is to test whether personality traits are associated with metabolic syndrome in a large...
Show moreThe prevalence of metabolic syndrome has paralleled the sharp increase in obesity. Given its tremendous physical, emotional, and financial burden, it is of critical importance to identify who is most at risk and the potential points of intervention. Psychological traits, in addition to physiological and social risk factors, may contribute to metabolic syndrome. The objective of the present research is to test whether personality traits are associated with metabolic syndrome in a large community sample. Participants (N = 5,662) from Sardinia, Italy, completed a comprehensive personality questionnaire, the NEO-PI-R, and were assessed on all components of metabolic syndrome (waist circumference, triglycerides, high-density lipoprotein cholesterol, blood pressure, and fasting glucose). Logistic regressions were used to predict metabolic syndrome from personality traits, controlling for age, sex, education, and current smoking status. Among adults over age 45 (n = 2,419), Neuroticism and low Agreeableness were associated with metabolic syndrome, whereas high Conscientiousness was protective. Individuals who scored in the top 10% on Conscientiousness were approximately 40% less likely to have metabolic syndrome (OR = 0.61, 95% CI = 0.41-0.92), whereas those who scored in the lowest 10% on Agreeableness were 50% more likely to have it (OR = 1.53, 95% CI = 1.09-2.16). At the facet level, traits related to impulsivity and hostility were the most strongly associated with metabolic syndrome. The present research indicates that those with fewer psychological resources are more vulnerable to metabolic syndrome and suggests a psychological component to other established risk factors.
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Date Issued
2010
Identifier
FSU_migr_mhs-0020, 10.1007/s11357-010-9153-9
Format
Citation
X-ray Structure and Biophysical Properties of Rabbit Fibroblast Growth Factor 1
Title
X-ray Structure and Biophysical Properties of Rabbit Fibroblast Growth Factor 1.
Creator
Lee, Jihun, Blaber, Sachiko, Irsigler, Andre, Aspinwall, Eric, Blaber, Michael
Abstract/Description
The rabbit is an important and de facto animal model in the study of ischemic disease and angiogenic therapy. Additionally, fibroblast growth factor 1 (FGF-1) is emerging as one of the most important growth factors for novel proangiogenic and pro-arteriogenic therapy. However, despite its significance, the fundamental biophysical properties of rabbit FGF-1, including its X-ray structure, have never been reported. Here, the cloning, crystallization, X-ray structure and determination of the...
Show moreThe rabbit is an important and de facto animal model in the study of ischemic disease and angiogenic therapy. Additionally, fibroblast growth factor 1 (FGF-1) is emerging as one of the most important growth factors for novel proangiogenic and pro-arteriogenic therapy. However, despite its significance, the fundamental biophysical properties of rabbit FGF-1, including its X-ray structure, have never been reported. Here, the cloning, crystallization, X-ray structure and determination of the biophysical properties of rabbit FGF-1 are described. The X-ray structure shows that the amino-acid differences between human and rabbit FGF-1 are solvent-exposed and therefore potentially immunogenic, while the biophysical studies identify differences in thermostability and receptor-binding affinity that distinguish rabbit FGF-1 from human FGF-1.
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Date Issued
2009
Identifier
FSU_migr_biomed_faculty_publications-0016, 10.1107/S1744309109040287, PMC2777034
Format
Citation
Engineering an Improved Crystal Contact Across a Solvent-Mediated Interface of Human Fibroblast Growth Factor 1
Title
Engineering an Improved Crystal Contact Across a Solvent-Mediated Interface of Human Fibroblast Growth Factor 1.
Creator
Meher, Akshaya, Blaber, Sachiko, Lee, Jihun, Honjo, Ejiro, Kuroki, Ryota, Blaber, Michael
Abstract/Description
Large-volume protein crystals are a prerequisite for neutron diffraction studies and their production represents a bottleneck in obtaining neutron structures. Many protein crystals that permit the collection of high-resolution X-ray diffraction data are inappropriate for neutron diffraction owing to a plate-type morphology that limits the crystal volume. Human fibroblast growth factor 1 crystallizes in a plate morphology that yields atomic resolution X-ray diffraction data but has...
Show moreLarge-volume protein crystals are a prerequisite for neutron diffraction studies and their production represents a bottleneck in obtaining neutron structures. Many protein crystals that permit the collection of high-resolution X-ray diffraction data are inappropriate for neutron diffraction owing to a plate-type morphology that limits the crystal volume. Human fibroblast growth factor 1 crystallizes in a plate morphology that yields atomic resolution X-ray diffraction data but has insufficient volume for neutron diffraction. The thin physical dimension has been identified as corresponding to the b cell edge and the X-ray structure identified a solvent-mediated crystal contact adjacent to position Glu81 that was hypothesized to limit efficient crystal growth in this dimension. In this report, a series of mutations at this crystal contact designed to both reduce side-chain entropy and replace the solvent-mediated interface with direct side-chain contacts are reported. The results suggest that improved crystal growth is achieved upon the introduction of direct crystal contacts, while little improvement is observed with side-chain entropy-reducing mutations alone.
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Date Issued
2009
Identifier
FSU_migr_biomed_faculty_publications-0017, 10.1107/S1744309109036987, PMC2777043
Format
Citation
Mobile Health Unit for Obesity Education
Title
Mobile Health Unit for Obesity Education: Attitudes and Perceptions Among Local Homeless Residents.
Creator
Soberon, Angelica, Brown, L., Rodríguez, Jose
Abstract/Description
Objective: Poverty is a risk factor for obesity. Using homelessness as a marker for poverty, we examined the prevalence of obesity in Leon County, Florida's homeless population to determine their attitudes and perceptions towards the disease. Methods: Our medical team equipped a mobile health unit with educational material before visiting homeless shelters, health fairs and shopping areas in Northwest Florida. Participants at these sites completed a short survey and our staff calculated Body...
Show moreObjective: Poverty is a risk factor for obesity. Using homelessness as a marker for poverty, we examined the prevalence of obesity in Leon County, Florida's homeless population to determine their attitudes and perceptions towards the disease. Methods: Our medical team equipped a mobile health unit with educational material before visiting homeless shelters, health fairs and shopping areas in Northwest Florida. Participants at these sites completed a short survey and our staff calculated Body Mass Index (BMI). The team also distributed lifestyle modification and obesity patient education. BMI and survey responses were compared and analyzed using statistical software. Results: Subjects were assigned to one of two groups: the homeless group or the "all others" group. The homeless group's responses and BMIs were compared to the "all others" group. Weight status in the homeless group was distributed as follows: normal weight (48.3%), overweight (39.7%) and obese (12.1%). The distribution among all other participants was: normal weight (34.8%), overweight (25.8%), and obese (39.3%). Homeless participants overestimated their BMI category as often as all others, were less likely to participate in weight loss interventions, and less likely to recognize overweight and obesity as a problem. Conclusion: Medical practitioners need to recognize the role of patients' attitudes about weight loss and clarify definitions of exercise. Obesity is less prevalent among the homeless; however, the same obesity trends exist. In addition, we learned that the homeless are eager to learn about obesity and healthy lifestyles, but may not be as willing to attempt interventions that have worked in other populations. Mobile health units can be used to reach out to them.
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Date Issued
2009
Identifier
FSU_migr_fmr-0035
Format
Citation
S(1)' and S(2)' Subsite Specificities of Human Plasma Kallikrein and Tissue Kallikrein 1 for the Hydrolysis of Peptides Derived from the Bradykinin Domain of Human Kininogen
Title
S(1)' and S(2)' Subsite Specificities of Human Plasma Kallikrein and Tissue Kallikrein 1 for the Hydrolysis of Peptides Derived from the Bradykinin Domain of Human Kininogen.
Creator
Lima, Aurelio, Alves, Fabiana, Angelo, Pedro, Andrade, Douglas, Blaber, Sachiko, Blaber, Michael, Juliano, Luiz, Juliano, Maria
Abstract/Description
The S(1)' and S(2)' subsite specificities of human tissue kallikrein 1 (KLK1) and human plasma kallikrein (HPK) were examined with the peptide series Abz-GFSPFRXSRIQ-EDDnp and Abz-GFSPFRSXRIQ-EDDnp [X=natural amino acids or S(PO(3)H(2))]. KLK1 efficiently hydrolyzed most of the peptides except those containing negatively charged amino acids at P(1)' and P(2)' positions. Abz-GFSPFRSSRIQ-EDDnp, as in human kininogen, is the best substrate for KLK1 and exclusively cleaved the R-S bond. All other...
Show moreThe S(1)' and S(2)' subsite specificities of human tissue kallikrein 1 (KLK1) and human plasma kallikrein (HPK) were examined with the peptide series Abz-GFSPFRXSRIQ-EDDnp and Abz-GFSPFRSXRIQ-EDDnp [X=natural amino acids or S(PO(3)H(2))]. KLK1 efficiently hydrolyzed most of the peptides except those containing negatively charged amino acids at P(1)' and P(2)' positions. Abz-GFSPFRSSRIQ-EDDnp, as in human kininogen, is the best substrate for KLK1 and exclusively cleaved the R-S bond. All other peptides were cleaved also at the F-R bond. The synthetic human kininogen segment Abz-MISLMKRPPGFSPFRS(390)S(391)RI-NH(2) was hydrolyzed by KLK1 first at R-S and then at M-K bonds, releasing Lys-bradykinin. In the S(390) and S(391) phosphorylated analogs, this order of hydrolysis was inverted due to the higher resistance of the R-S bond. Abz-MISLMKRPPG-FSPFRSS(PO(3)H(2))(391)RI-NH(2) was hydrolyzed by KLK1 at M-K and mainly at the F-R bond, releasing des-(Arg(9))-Lys-Bk which is a B1 receptor agonist. HPK cleaved all the peptides at R and showed restricted specificity for S in the S(1)' subsite, with lower specificity for the S(2)' subsite. Abz-MISLMKRPPGFSPFRSSRI-NH(2) was efficiently hydrolyzed by HPK under bradykinin release, while the analogs containing S(PO(3)H(2)) were poorly hydrolyzed. In conclusion, S(1)' and S(2)' subsite specificities of KLK1 and HPK showed peculiarities that were observed with substrates containing the amino acid sequence of human kininogen.
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Date Issued
2008
Identifier
FSU_migr_biomed_faculty_publications-0011
Format
Citation
Protease-Activated Receptor Dependent and Independent Signaling by Kallikreins 1 and 6 in CNS Neuron and Astroglial Cell Lines
Title
Protease-Activated Receptor Dependent and Independent Signaling by Kallikreins 1 and 6 in CNS Neuron and Astroglial Cell Lines.
Creator
Vandell, Alexander, Larson, Nadya, Laxmikanthan, Gurunathan, Panos, Michael, Blaber, Sachiko, Blaber, Michael, Scarisbrick, Isobel
Abstract/Description
While protease-activated receptors (PARs) are known to mediate signaling events in CNS, contributing both to normal function and pathogenesis, the endogenous activators of CNS PARs are poorly characterized. In this study, we test the hypothesis that kallikreins (KLKs) represent an important pool of endogenous activators of CNS PARs. Specifically, KLK1 and KLK6 were examined for their ability to evoke intracellular Ca(2+) flux in a PAR-dependent fashion in NSC34 neurons and Neu7 astrocytes....
Show moreWhile protease-activated receptors (PARs) are known to mediate signaling events in CNS, contributing both to normal function and pathogenesis, the endogenous activators of CNS PARs are poorly characterized. In this study, we test the hypothesis that kallikreins (KLKs) represent an important pool of endogenous activators of CNS PARs. Specifically, KLK1 and KLK6 were examined for their ability to evoke intracellular Ca(2+) flux in a PAR-dependent fashion in NSC34 neurons and Neu7 astrocytes. Both KLKs were also examined for their ability to activate mitogen-activated protein kinases (extracellular signal-regulated kinases, C-Jun N-terminal kinases, and p38) and protein kinase B (AKT) intracellular signaling cascades. Cumulatively, these studies show that KLK6, but not KLK1, signals through PARs. KLK6 evoked intracellular Ca(2+) flux was mediated by PAR1 in neurons and both PAR1 and PAR2 in astrocytes. Importantly, both KLK1 and KLK6 altered the activation state of mitogen-activated protein kinases and AKT, suggestive of important roles for each in CNS neuron and glial differentiation, and survival. The cellular specificity of CNS-KLK activity was underscored by observations that both proteases promoted AKT activation in astrocytes, but inhibited such signaling in neurons. PAR1 and bradykinin receptor inhibitors were used to demonstrate that KLK1-mediated activation of extracellular signal-regulated kinases in neurons occurred in a non-PAR, bradykinin 2 (B2) receptor-dependent fashion, while similar signaling by KLK6 was mediated by the combined activation of PAR1 and B2. Cumulatively results indicate KLK6, but not KLK1 is an activator of CNS PARs, and that both KLKs are poised to signal in a B2 receptor-dependent fashion to regulate multiple signal transduction pathways relevant to CNS physiologic function and dysfunction.
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Date Issued
2008
Identifier
FSU_migr_biomed_faculty_publications-0010
Format
Citation
Substrate Specificity of Human Kallikreins 1 and 6 Determined by Phage Display
Title
Substrate Specificity of Human Kallikreins 1 and 6 Determined by Phage Display.
Creator
Li, Hai-Xin, Hwang, Bum-Yeol, Laxmikanthan, Gurunathan, Blaber, Sachiko, Blaber, Michael, Golubkov, Pavel, Ren, Pengyu, Iverson, Brent, Georgiou, George
Abstract/Description
The human tissue kallikrein (KLK) family contains 15 secreted serine proteases that are expressed in a wide range of tissues and have been implicated in different physiological functions and disease states. Of these, KLK1 has been shown to be involved in the regulation of multiple physiological processes such as blood pressure, smooth muscle contraction, and vascular cell growth. KLK6 is overexpressed in breast and ovarian cancer tissues and has been shown to cleave peptide derived from human...
Show moreThe human tissue kallikrein (KLK) family contains 15 secreted serine proteases that are expressed in a wide range of tissues and have been implicated in different physiological functions and disease states. Of these, KLK1 has been shown to be involved in the regulation of multiple physiological processes such as blood pressure, smooth muscle contraction, and vascular cell growth. KLK6 is overexpressed in breast and ovarian cancer tissues and has been shown to cleave peptide derived from human myelin protein and Abeta amyloid peptide in vitro. Here we analyzed the substrate specificity of KLK1 and KLK6, by substrate phage display using a random octapeptide library. Consistent with earlier biochemical data, KLK1 was shown to exhibit both trypsin- and chymotrypsin-like selectivities with Tyr/Arg preferred at site P1, Ser/Arg strongly preferred at P1', and Phe/Leu at P2. KLK6 displayed trypsin-like activity, with the P1 position occupied only by Arg and a strong preference for Ser in P1'. Docking simulations of consensus peptide provide information on the identity of the enzyme residues that are responsible for substrate binding. Bioinformatic analysis suggested several putative KLK6 protein substrates, such as ionotropic glutamate receptor (GluR) and synphilin.
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Date Issued
2008
Identifier
FSU_migr_biomed_faculty_publications-0006
Format
Citation
Kallikreins are Associated with Secondary Progressive Multiple Sclerosis and Promote Neurodegeneration
Title
Kallikreins are Associated with Secondary Progressive Multiple Sclerosis and Promote Neurodegeneration.
Creator
Scarisbrick, Isobel, Linbo, Rachel, Vandell, Alexander, Keegan, Mark, Blaber, Sachiko, Blaber, Michael, Sneve, Diane, Lucchinetti, Claudia F., Rodriguez, Moses, Diamandis,...
Show moreScarisbrick, Isobel, Linbo, Rachel, Vandell, Alexander, Keegan, Mark, Blaber, Sachiko, Blaber, Michael, Sneve, Diane, Lucchinetti, Claudia F., Rodriguez, Moses, Diamandis, Eleftherios P.
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Abstract/Description
Tissue kallikrein KLK1 and the kallikrein-related peptidases KLK2-15 are a subfamily of serine proteases that have defined or proposed roles in a range of central nervous system (CNS) and non-CNS pathologies. To further understand their potential activity in multiple sclerosis (MS), serum levels of KLK1, 6, 7, 8 and 10 were determined in 35 MS patients and 62 controls by quantitative fluorometric ELISA. Serum levels were then correlated with Expanded Disability Status Scale (EDSS) scores...
Show moreTissue kallikrein KLK1 and the kallikrein-related peptidases KLK2-15 are a subfamily of serine proteases that have defined or proposed roles in a range of central nervous system (CNS) and non-CNS pathologies. To further understand their potential activity in multiple sclerosis (MS), serum levels of KLK1, 6, 7, 8 and 10 were determined in 35 MS patients and 62 controls by quantitative fluorometric ELISA. Serum levels were then correlated with Expanded Disability Status Scale (EDSS) scores determined at the time of serological sampling or at last clinical follow-up. Serum levels of KLK1 and KLK6 were elevated in MS patients (p
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Date Issued
2008
Identifier
FSU_migr_biomed_faculty_publications-0008
Format
Citation
Mutagenesis of the Crystal Contact of Acidic Fibroblast Growth Factor
Title
Mutagenesis of the Crystal Contact of Acidic Fibroblast Growth Factor.
Creator
Honjo, Eijiro, Tamada, Taro, Adachi, Motoyasu, Kuroki, Ryota, Meher, Akshaya, Blaber, Michael
Abstract/Description
An attempt has been made to improve a crystal contact of human acidic fibroblast growth factor (haFGF; 140 amino acids) to control the crystal growth, because haFGF crystallizes only as a thin-plate form, yielding crystals suitable for X-ray but not neutron diffraction. X-ray crystal analysis of haFGF showed that the Glu81 side chain, located at a crystal contact between haFGF molecules, is in close proximity with an identical residue related by crystallographic symmetry, suggesting that...
Show moreAn attempt has been made to improve a crystal contact of human acidic fibroblast growth factor (haFGF; 140 amino acids) to control the crystal growth, because haFGF crystallizes only as a thin-plate form, yielding crystals suitable for X-ray but not neutron diffraction. X-ray crystal analysis of haFGF showed that the Glu81 side chain, located at a crystal contact between haFGF molecules, is in close proximity with an identical residue related by crystallographic symmetry, suggesting that charge repulsion may disrupt suitable crystal-packing interactions. To investigate whether the Glu residue affects the crystal-packing interactions, haFGF mutants in which Glu81 was replaced by Ala, Val, Leu, Ser and Thr were constructed. Although crystals of the Ala and Leu mutants were grown as a thin-plate form by the same precipitant (formate) as the wild type, crystals of the Ser and Thr mutants were grown with increased thickness, yielding a larger overall crystal volume. X-ray structural analysis of the Ser mutant determined at 1.35 A resolution revealed that the hydroxy groups of Ser are linked by hydrogen bonds mediated by the formate used as a precipitant. This approach to engineering crystal contacts may contribute to the development of large protein crystals for neutron crystallography.
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Date Issued
2008
Identifier
FSU_migr_biomed_faculty_publications-0007
Format
Citation