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B Brodsky

Publications and source records attributed to B Brodsky.

At least 19 recordsLinked to original sources
PubMed2001

The crystal and molecular structure of a collagen-like peptide with a biologically relevant sequence.

A detailed description of the 2.0 A structure of the triple-helical peptide, (Pro-Hyp-Gly)(3)-Ile-Thr-Gly-Ala-Arg-Gly-Leu-Ala-Gly-Pro-Hyp-Gly-(Pro-Hyp-Gly)(3), denoted as T3-785, is presented. This peptide contains a biologically relevant sequence and was designed to model the imino acid-poor 785-796 region of human type III collagen just C-terminal to the matrix metalloproteinase cleavage site. The crystal structure of the T3-785 peptide demonstrates that sequence can influence local conformational changes in triple-helical structure, in terms of superhelical pitch, hydrogen bonding pattern, and hydration patterns. The novel packing arrangement displayed by the T3-785 structure, compared with those of collagen-like peptides with more imino acid-rich sequences indicates the sequence dependence of intermolecular assemblies in collagen as well. The observed synergy between the packing arrangements and the extended hydration network indicates that hydration of the triple helix is directly related to its association with other molecules.

Amino Acid Motifs↗
PubMed2001

Effects of iodine on inducible nitric oxide synthase and cyclooxygenase-2 expression in sulfur mustard-induced skin.

In a previous study we demonstrated the protective effect of topical iodine as postexposure treatment for sulfur mustard (SM) application. The iodine treatment results in significantly reduced inflammation and necrosis and increased epidermal hyperplasia. The expression and localization of inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2) in paraffin-embedded skin samples from that study were evaluated in the present investigation. We compared the immunoreactivity of iNOS and COX-2 using five samples from each of the following four test sites: untreated control sites, SM-exposed sites, sites treated with iodine mixture 15 min after SM exposure, and sites treated with iodine 30 min after SM exposure. All animals were killed 2 days after irritant exposure. iNOS immunoreactivity was present only in skin sites exposed to SM without iodine treatment. The ulcerated skin was covered with a relatively thick band of exudate composed of iNOS-immunostained polymorphonuclear cells and macrophages. In untreated skin, COX-2 immunostaining was limited to the thin suprabasal epidermal layer. In SM-exposed skin, induction of COX-2 was noted in inflammatory cells located close to the site of epidermal injury. In skin sites treated with iodine 15 or 30 min after SM exposure, the regenerating hyperplastic epithelium showed moderate cytoplasmic staining localized to the epithelium overlying the basal layer. This pattern of staining was also present in the nearby dermal fibroblasts. Thus, in contrast to the skin samples exposed to SM without iodine treatment, the epidermal layer expressing immunohistochemical positivity for COX-2 was thicker and corresponded to the epidermal hyperplasia noted in samples treated with iodine. It is well documented that prostaglandins (PGs) promote epidermal proliferation, thereby contributing to the repair of injured skin. That the induction of the COX-2 shown in our study may also play a role in the healing process is indicated by the present evidence. The results suggest that nitric oxide radicals (NO*) are involved in mediating the damage induced by the SM and that iodine-related reduction in acute epidermal inflammation is associated with reduced iNOS expression.

Administration, Topical↗
PubMed2000

Interaction of collagen-like peptide models of asymmetric acetylcholinesterase with glycosaminoglycans: spectroscopic studies of conformational changes and stability.

The effect of heparin on the conformation and stability of triple-helical peptide models of the collagen tail of asymmetric acetylcholinesterase expands our understanding of heparin interactions with proteins and presents an opportunity for clarifying the nature of binding of ligands to collagen triple-helix domains. Within the collagen tail of AChE, there are two consensus sequences for heparin binding of the form BBXB, surrounded by additional basic residues. Circular dichroism studies were used to determine the effect of the addition of increasing concentrations of heparin on triple-helical peptide models for the heparin binding domains, including peptides in which the basic residues within and surrounding the consensus sequence were replaced by alanine residues. The addition of heparin caused an increased triple-helix content with saturation properties for the peptide modeling the C-terminal site, while precipitation, with no increased helix content resulted from heparin addition to the peptide modeling the N-terminal site. The results suggest that the two binding sites with a similar triple-helical conformation have distinctive ways of interacting with heparin, which must relate to small differences in the consensus sequence (GRKGR vs GKRGK) and in the surrounding basic residues. Addition of heparin increased the thermal stability of all peptides containing the consensus sequence. Heparan sulfate produced conformational and stabilization effects similar to those of heparin, while chondroitin sulfate led to a cloudy solution, loss of circular dichroism signal, and a smaller increase in thermal stability. Thus, specificity in both the sequence of the triple helix and the type of glycosaminoglycan is required for this interaction.

Acetylcholinesterase↗
PubMed2000

Amino acid propensities for the collagen triple-helix.

Determination of the tendencies of amino acids to form alpha-helical and beta-sheet structures has been important in clarifying stabilizing interactions, protein design, and the protein folding problem. In this study, we have determined for the first time a complete scale of amino acid propensities for another important protein motif: the collagen triple-helix conformation with its Gly-X-Y repeating sequence. Guest triplets of the form Gly-X-Hyp and Gly-Pro-Y are used to quantitate the conformational propensities of all 20 amino acids for the X and Y positions in the context of a (Gly-Pro-Hyp)(8) host peptide. The rankings for both the X and Y positions show the highly stabilizing nature of imino acids and the destabilizing effects of Gly and aromatic residues. Many residues show differing propensities in the X versus Y position, related to the nonequivalence of these positions in terms of interchain interactions and solvent exposure. The propensity of amino acids to adopt a polyproline II-like conformation plays a role in their triple-helix rankings, as shown by a moderate correlation of triple-helix propensity with frequency of occurrence in polyproline II-like regions. The high propensity of ionizable residues in the X position suggests the importance of interchain hydrogen bonding directly or through water to backbone carbonyls or hydroxyprolines. The low propensity of side chains with branching at the C(delta) in the Y position supports models suggesting these groups block solvent access to backbone C=O groups. These data provide a first step in defining sequence-dependent variations in local triple-helix stability and binding, and are important for a general understanding of side chain interactions in all proteins.

Amino Acids↗
PubMed2000

Topical iodine preparation as therapy against sulfur mustard-induced skin lesions.

Sulfur mustard (SM) is a powerful vesicant employed as an agent of chemical warfare. This study demonstrates the therapeutic effect of a novel topical iodine preparation as a postexposure treatment against SM-induced lesions in the fur-covered guinea-pig skin model. Iodine treatment 15 min after SM exposure resulted in statistically significant reductions of 48, 50, and 55% in dermal acute inflammation, hemorrhage, and necrosis, respectively, whereas, the epidermal healing markers, hyperkerathosis and acanthosis, were significantly elevated by 72 and 67%, respectively, 2 days after treatment. At the interval of 30 min between SM exposure and iodine treatment, there was a significant degree of healing or recovery, albeit to a lesser extent than that observed in the shorter interval. Although the epidermal healing markers were not elevated, the parameters indicative of active tissue damage, such as subepidermal microblisters, epidermal ulceration, dermal acute inflammation, hemorrhage, and necrosis, were significantly reduced by 35, 67, 43, 39, and 45%, respectively. At the 45-min interval between exposure and treatment, there was also a certain degree of healing or recovery expressed as significant reductions in dermal subacute inflammation, subepidermal microblister formation, and epidermal ulceration, whereas, acanthosis was statistically elevated, indicating an increased healing potential. At the 60-min interval, iodine was less efficacious; nevertheless, a significant reduction in the incidence of subepidermal microblisters and an expansion of the acanthotic area were observed. Gross ulceration was significantly decreased at intervals of 15 and 30 min between exposure and treatment. The local anesthetic, lidocaine, did not alter the therapeutic effect of iodine. SM was not affected chemically by iodine as measured by gas chromatography-mass spectrometry (GC-MS) analysis. These findings suggest that the iodine preparation functions as an antidote against skin lesions induced by SM.

Administration, Topical↗
PubMed2000

Staggered molecular packing in crystals of a collagen-like peptide with a single charged pair.

The crystal structure of the triple-helical peptide, (Pro-Hyp-Gly)(4)-Glu-Lys-Gly-(Pro-Hyp-Gly)(5) has been determined to 1.75 A resolution. This peptide was designed to examine the effect of a pair of adjacent, oppositely charged residues on collagen triple-helical conformation and intermolecular interactions. The molecular conformation (a 7(5) triple helix) and hydrogen bonding schemes are similar to those previously reported for collagen triple helices and provides a second instance of water mediated N--H . . . O==C interchain hydrogen bonds for the amide group of the residue following Gly. Although stereochemically capable of forming intramolecular or intermolecular ion pairs, the lysine and glutamic acid side-chains instead display direct interactions with carbonyl groups and hydroxyproline hydroxyl groups or interactions mediated by water molecules. Solution studies on the EKG peptide indicate stabilization at neutral pH values, where both Glu and Lys are ionized, but suggest that this occurs because of the effects of ionization on the individual residues, rather than ion pair formation. The EKG structure suggests a molecular mechanism for such stabilization through indirect hydrogen bonding. The molecular packing in the crystal includes an axial stagger between molecules, reminiscent of that observed in D-periodic collagen fibrils. The presence of a Glu-Lys-Gly triplet in the middle of the sequence appears to mediate this staggered molecular packing through its indirect water-mediated interactions with backbone C==O groups and side chains.

Amino Acid Sequence↗
PubMed2000

Interaction of the collagen-like tail of asymmetric acetylcholinesterase with heparin depends on triple-helical conformation, sequence and stability.

The collagen-like tail of asymmetric acetylcholinesterase (AChE) contains two heparin-binding domains (HBDs) that interact with heparan sulphate proteoglycans, determining the anchoring of the enzyme at the basal lamina and its specific localization at the neuromuscular junction. Both HBDs are characterized by a cluster of basic residues containing a core with the BBXB consensus sequence (where B represents a basic residue and X a non-basic residue). To study the interaction of such HBDs with heparin we have used synthetic peptides to model the N-terminal and C-terminal sites. CD spectroscopy showed that all peptides are triple-helical at low temperatures, and undergo trimer-to-monomer transitions. Displacement assays of asymmetric AChE bound to heparin were performed using the peptides in both monomeric and triple-helical states. In the monomeric conformation, all the peptides were able to displace low levels of AChE depending on the basic charge content. In the triple-helical conformation, peptides containing the consensus sequence showed a large increase in the ability to displace bound AChE. Results suggest that the specific binding of the collagen-like-tail peptides to heparin depends both on the presence of the core sequence and on the triple-helical conformation. Moreover, BBXB-containing peptides that are less stable are more effective in displacing AChE, suggesting that the interaction region needs a significant amount of structural flexibility to better accommodate the ligand.

Acetylcholinesterase↗
PubMed2000

Site-specific NMR monitoring of cis-trans isomerization in the folding of the proline-rich collagen triple helix.

Understanding the folding of the proline-rich collagen triple helix requires consideration of the effects of proline cis-trans isomerization and may shed light on the misfolding of collagen in connective tissue diseases. Folding was monitored in real time by heteronuclear 2D NMR spectroscopy for the (15)N labeled positions in the triple-helical peptide T1-892 [GPAGPAGPVGPAGARGPAGPOGPOGPOGPOGV]. In the equilibrium unfolded monomer form, each labeled residue showed multiple peaks with interconversion rates consistent with cis-trans isomerization of Gly-Pro and Pro-Hyp bonds. Real-time NMR studies on the folding of T1-892 showed slow decay of monomer peaks and a concomitant increase in trimer peaks. Gly25 in the C-terminal rich (Gly-Pro-Hyp)(4) domain folds first, consistent with its being a nucleation domain. Analysis of the kinetics indicates that the folding of Gly25 is biphasic and the slower step represents cis-trans isomerization of imino acids. This illustrates that nucleation is limited by cis-trans isomerization. Monitoring Gly6, Gly10, Ala12, and Gly13 monomer and trimer peaks captures the C- to N-terminal propagation of the triple helix, which is also limited by Gly-Pro cis-trans isomerization events. The zipper-like nature of the propagation process is confirmed by the slower rate of folding of Ala6 compared to Gly13, reflecting the larger number of isomerization events encountered by the more N-terminal Ala6. The cis-trans isomerization events at multiple proline residues is a complex statistical process which can be visualized by these NMR studies.

Amino Acid Sequence↗
PubMed2000

Destabilization of osteogenesis imperfecta collagen-like model peptides correlates with the identity of the residue replacing glycine.

Mutations resulting in replacement of one obligate Gly residue within the repeating (Gly-Xaa-Yaa)(n) triplet pattern of the collagen type I triple helix are the major cause of osteogenesis imperfecta (OI). Phenotypes of OI involve fragile bones and range from mild to perinatal lethal. In this study, host-guest triple-helical peptides of the form acetyl-(Gly-Pro-Hyp)(3)-Zaa-Pro-Hyp-(Gly-Pro-Hyp)(4)-Gly-Gly-amide are used to isolate the influence of the residue replacing Gly on triple-helix stability, with Zaa = Gly, Ala, Arg, Asp, Glu, Cys, Ser, or Val. Any substitution for Zaa = Gly (melting temperature, T(m) = 45 degrees C) results in a dramatic destabilization of the triple helix. For Ala and Ser, T(m) decreases to approximately 10 degrees C, and for the Arg-, Val-, Glu-, and Asp-containing peptides, T(m) < 0 degrees C. A Gly --> Cys replacement results in T(m) < 0 degrees C under reducing conditions but shows a broad transition (T(m) approximately 19 degrees C) in an oxidizing environment. Addition of trimethylamine N-oxide increases T(m) by approximately 5 degrees C per 1 M trimethylamine N-oxide, resulting in stable triple-helix formation for all peptides and allowing comparison of relative stabilities. The order of disruption of different Gly replacements in these peptides can be represented as Ala </= Ser < CPO(red) < Arg < Val < Glu </= Asp. The rank of destabilization of substitutions for Gly in these Gly-Pro-Hyp-rich homotrimeric peptides shows a significant correlation with the severity of natural OI mutations in the alpha1 chain of type I collagen.

Amino Acid Substitution↗
PubMed2000

Collagen model peptides: Sequence dependence of triple-helix stability.

The triple helix is a specialized protein motif, found in all collagens as well as in noncollagenous proteins involved in host defense. Peptides will adopt a triple-helical conformation if the sequence contains its characteristic features of Gly as every third residue and a high content of Pro and Hyp residues. Such model peptides have proved amenable to structural studies by x-ray crystallography and NMR spectroscopy, suitable for thermodynamic and kinetic analysis, and a valuable tool in characterizing the binding activities of the collagen triple helix. A systematic approach to understanding the amino acid sequence dependence of the collagen triple helix has been initiated, based on a set of host-guest peptides of the form, (Gly-Pro-Hyp)(3)-Gly-X-Y-(Gly-Pro-Hyp)(4). Comparison of their thermal stabilities has led to a propensity scale for the X and Y positions, and the additivity of contributions of individual residues is now under investigation. The local and global stability of the collagen triple helix is normally modulated by the residues in the X and Y positions, with every third position occupied by Gly in fibril-forming collagens. However, in collagen diseases, such as osteogenesis imperfecta, a single Gly may be substituted by another residue. Host-guest studies where the Gly is replaced by various amino acids suggest that the identity of the residue in the Gly position affects the degree of destabilization and the clinical severity of the disease.

Amino Acid Motifs↗
PubMed2000

Protective effect of povidone iodine ointment against skin lesions induced by chemical and thermal stimuli.

Mustard gas (sulfur mustard, HD) is a powerful vesicant employed as a chemical weapon. The present study demonstrates the effect of povidone iodine (PI) ointment against skin toxicity caused by HD. Gross and histopathological examinations showed that application of PI 20 min or less following exposure to the vesicant resulted in marked skin protection. The shorter, interval between exposure and treatment, the better was the protection achieved. Povidone iodine was also effective against other mustards, such as carboxybutylchloroethyl sulfide (CBCS) and mechlorethamine. The fact that PI protected the skin against agents that cannot be oxidized, such as iodoacetic acid, divinylsulfone and cantharidine, indicated that the antidotal effect of PI was unrelated to oxidation of the nitrogen and sulfur atoms of the mustards. Furthermore, NMR spectroscopy of CBCS treated with iodine did not show oxidation of the sulfur atom. Clinical experience with patients after accidential heat burns (mostly of grade I) has shown that topical application of PI ointment immediately after the stimulus significantly reduced, and often prevented, skin lesions. Apart from being a safe and widely used disinfectant, PI ointment is recommended as an efficient protective agent against skin toxicity caused by hazardous chemicals and by heat stimuli.

Administration, Topical↗
PubMed2000

Suicidal behavior in bipolar mood disorder: clinical characteristics of attempters and nonattempters.

OBJECTIVE: Bipolar Disorder is associated with a higher frequency of attempted suicide than most other psychiatric disorders. The reasons are unknown. This study compared bipolar subjects with a history of a suicide attempt to those without such a history, assessing suicidal behavior qualitatively and quantitatively, and examining possible demographic, psychopathologic and familial risk factors. METHODS: Patients (ages 18 to 75) with a DSM III-R Bipolar Disorder (n = 44) diagnosis determined by a structured interview for Axis I disorders were enrolled. Acute psychopathology, hopelessness, protective factors, and traits of aggression and impulsivity were measured. The number, method and degree of medical damage was assessed for suicide attempts, life-time. RESULTS: Bipolar suicide attempters had more life-time episodes of major depression, and twice as many were in a current depressive or mixed episode, compared to bipolar nonattempters. Attempters reported more suicidal ideation immediately prior to admission, and fewer reasons for living even when the most recent suicide attempt preceded the index hospitalization by more than six months. Attempters had more lifetime aggression and were more likely to be male. However, attempters did not differ from nonattempters on lifetime impulsivity. LIMITATIONS: The generalizability of the results is limited because this is a study of inpatients with a history of suicide attempts. Patients with Bipolar I and NOS Disorders were pooled and a larger sample is needed to look at differences. We could not assess psychopathology immediately prior to the suicide attempt because, only half of the suicide attempters had made attempts in the six months prior to admission. Patients with current comorbid substance abuse were excluded. No suicide completers were studied. CONCLUSIONS: Bipolar subjects with a history of suicide attempt experience more episodes of depression, and react to them by having severe suicidal ideation. Their diathesis for acting on feelings of anger or suicidal ideation is suggested by a higher level of lifetime aggression and a pattern of repeated suicide attempts.

Adult↗
PubMed2000

Anxiety in major depression: relationship to suicide attempts.

OBJECTIVE: This study was an examination of the relationship of lifetime panic disorder and anxiety symptoms at index hospitalization to a history of a suicide attempt in patients with a major depressive episode. METHOD: A total of 272 inpatients with at least one major depressive episode, with or without a history of a suicide attempt, were entered into the study. They were given structured diagnostic interviews for axis I and axis II disorders. Suicide attempt history, current psychopathology, and traits of aggression and impulsivity were also assessed. RESULTS: The rates of panic disorder did not differ in the suicide attempters and nonattempters. Agitation, psychic anxiety, and hypochondriasis were more severe in the nonattempter group. A multivariate analysis confirmed that this effect was independent of aggression and impulsivity. CONCLUSIONS: Comorbid panic disorder in patients with major depression does not seem to increase the risk for lifetime suicide attempt. The presence of greater anxiety in the nonattempters warrants further investigation.

Adolescent↗
PubMed1999

Sequence dependence of the folding of collagen-like peptides. Single amino acids affect the rate of triple-helix nucleation.

The refolding of thermally denatured model collagen-like peptides was studied for a set of 21 guest triplets embedded in a common host framework: acetyl-(Gly-Pro-Hyp)3-Gly-Xaa-Yaa-(Gly-Pro-Hyp)4-Gly-Gly-amide. The results show a strong dependence of the folding rate on the identity of the guest Gly-Xaa-Yaa triplet, with the half-times for refolding varying from 6 to 110 min (concentration = 1 mg/ml). All triplets of the form Gly-Xaa-Hyp promoted rapid folding, with the rate only marginally dependent on the residue in the Xaa position. In contrast, triplets of the form Gly-Pro-Yaa and Gly-Xaa-Yaa were slower and showed a wide range of half-times, varying with the identity of the residues in the triplet. At low concentrations, the folding can be described by third-order kinetics, suggesting nucleation is rate-limiting. Data on the relative nucleation ability of different Gly-Xaa-Yaa triplets support the favorable nature of imino acids, the importance of hydroxyproline, the varying effects of the same residue in the Xaa position versus the Yaa position, and the difficulties encountered when leucine or aspartic acid are in the Yaa position. Information on the relative propensities of different tripeptide sequences to promote nucleation of the triple-helix in peptides will aid in identification of nucleation sites in collagen sequences.

Circular Dichroism↗
PubMed1999

Structural consequences of D-amino acids in collagen triple-helical peptides.

The effects of racemization of aspartic acid on triple-helical formation have been studied using a "host-guest" peptide approach where selected guest Gly-Xaa-Yaa triplets were included within a common acetyl-(Gly-Pro-Hyp)3-Gly-Xaa-Yaa-(Gly-Pro-Hyp)4-Gly-Gly-amide frame-work. Four guest triplets, Gly-Asp-Hyp and Gly-Asp-Ala where Asp is either L-Asp or D-Asp were studied. Thermal stability data indicated that incorporation of D-Asp residues prevented triple-helix formation in phosphate buffered saline, although triple-helical structures were formed in a stabilizing solvent, 67% aqueous ethylene glycol. In this solvent the melting temperatures of D-Asp containing peptides were more than 30 degrees C lower than the corresponding peptides containing L-Asp. For Gly-Asp-Ala peptides, but not Gly-Asp-Hyp, peptides, melting profiles indicated that a mixture of the D- and L-Asp containing peptides were able to form heterotrimer triple-helical molecules. These studies illustrate the dramatic destabilizing effect of D-amino acids on the triple-helix stability, but indicate that they can be accommodated in this conformation.

Amino Acid Sequence↗
PubMed1999

Folding of peptide models of collagen and misfolding in disease.

The misfolding of the triple helix has been shown to play a critical role in collagen diseases. Normal and mutated collagen triple helices can be modeled by short, synthetic peptides of varying design. NMR spectroscopy and circular dichroism studies on the assembly of these peptide models have recently been used to isolate specific steps in the folding pathway and have provided information on the alterations resulting from mutations.

Amino Acid Sequence↗
PubMed1998

Nuclear magnetic resonance shows asymmetric loss of triple helix in peptides modeling a collagen mutation in brittle bone disease.

To investigate a human folding disease, NMR studies were carried out on collagen-like peptides to define the structural consequences of a single amino acid change found in patients with osteogenesis imperfecta (OI), a disease characterized by fragile bones. One peptide included a normal collagen sequence, while a second peptide included a Gly --> Ser substitution as found in a nonlethal case of OI. Residue specific internal dynamics and conformational studies indicate that the normal collagen-like sequence forms a triple helix which is rigid along its entire length. The introduction of a Gly --> Ser substitution induces an asymmetric disruption of the uniform triple helix. While the C-terminal end of the peptide retains the triple helix, the Ser substitution site and residues N-terminal to it exhibit the mobility of a random chain. This equilibrium state indicates that a Gly substitution can terminate the C to N propagation of the triple helix and suggests that renucleation is required for folding to continue. Defective folding has been implicated in brittle bone disease, and these results begin to characterize the folding process in OI collagens. OI collagen studies may also provide insights about defective protein folding, assembly, and aggregation in other human diseases.

Amino Acid Sequence↗