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V P Marshall

Publications and source records attributed to V P Marshall.

At least 19 recordsLinked to original sources
PubMed1999

Thermodynamic and circular dichroism studies differentiate inhibitor interactions with the stromelysin S(1)-S(3) and S(')(1)-S(')(3) subsites.

Interactions of stromelysin with a series of inhibitors representative of three chemical templates with distinct binding modes were examined. Unfolding temperatures for inhibitor complexes were 10 degrees C to 15 degrees C greater than for apo stromelysin. Minor changes in ellipticity in the far-UV CD spectra of complexes indicated that ligand-induced conformational changes were localized to the binding site and did not involve gross changes in protein folding. Isothermal titrating calorimetry of thiadiazole-containing inhibitors, which bind in the S(1)-S(3) subsites of stromelysin, indicated that the binding interaction was exothermic and only slightly favorable entropically. Near-UV CD spectra showed large positive ellipticity increases from 250 to 300 nm, consistent with an interaction between the benzene ring of the inhibitor and stromelysin residues Tyr155 and Tyr168. Interactions between stromelysin and amide-hydroxamate ligands, which bind in the S(')(1)-S(')(3) subsites, were found to be both enthalpically and entropically driven. Binding of this class of ligands resulted in modest negative ellipticity changes at 260-285 nm and positive increases at 292 nm. Stromelysin complexed to a lactam-hydroxamate inhibitor with structure extending into both the S(1)-S(3) and S(')(1)-S(')(3) subsites showed increased ellipticity at 245 nm and negative changes at 260-285 and 295 nm.

Binding Sites↗
PubMed1999

Dynamics of stromelysin/inhibitor interactions studied by 15N NMR relaxation measurements: comparison of ligand binding to the S1-S3 and S'1-S'3 subsites.

This report describes the backbone amide dynamics of the uniformly 15N labeled catalytic domain of human stromelysin complexed to PNU-99533, a hydroxamate-containing ligand that binds to the S'1-S'3 region (right side) of the stromelysin active site, and to PNU-107859 and PNU-142372, both thiadiazole-containing ligands that bind to the S1-S3 region (left side) of the stromelysin active site. 15N R1, R2 and NOE NMR relaxation measurements were recorded and analyzed for each complex. Different dynamic behaviors were observed for stromelysin complexed to the two classes of ligands, indicating that it may be possible to use protein dynamics to distinguish between different binding orientations. In the absence of bound ligand at the S1-S3 subsites, the S1-S3 residues were found to be relatively rigid. In contrast, the S'1-S'3 subsites were found to be flexible in the absence of interactions with ligand. The relative rigidness of the S1-S3 subsites may be responsible for MMP binding specificity by discriminating between ligands of different shapes. By contrast, the inherent flexibility of the S'1-S'3 subsites allows structural rearrangement to accommodate a broad range of incoming substrates or inhibitors. Similarities and differences in dynamics observed for each complex provide insights into the interactions responsible for protein-ligand recognition. The relevance of protein dynamics to structure-based drug design is discussed.

Binding Sites↗
PubMed1998

Structural characterizations of nonpeptidic thiadiazole inhibitors of matrix metalloproteinases reveal the basis for stromelysin selectivity.

The binding of two 5-substituted-1,3,4-thiadiazole-2-thione inhibitors to the matrix metalloproteinase stromelysin (MMP-3) have been characterized by protein crystallography. Both inhibitors coordinate to the catalytic zinc cation via an exocyclic sulfur and lay in an unusual position across the unprimed (P1-P3) side of the proteinase active site. Nitrogen atoms in the thiadiazole moiety make specific hydrogen bond interactions with enzyme structural elements that are conserved across all enzymes in the matrix metalloproteinase class. Strong hydrophobic interactions between the inhibitors and the side chain of tyrosine-155 appear to be responsible for the very high selectivity of these inhibitors for stromelysin. In these enzyme/inhibitor complexes, the S1' enzyme subsite is unoccupied. A conformational rearrangement of the catalytic domain occurs that reveals an inherent flexibility of the substrate binding region leading to speculation about a possible mechanism for modulation of stromelysin activity and selectivity.

Animals↗
PubMed1998

Solution structures of stromelysin complexed to thiadiazole inhibitors.

Unregulated or overexpressed matrix metalloproteinases (MMPs), including stromelysin, collagenase, and gelatinase. have been implicated in several pathological conditions including arthritis and cancer. Small-molecule MMP inhibitors may have therapeutic value in the treatment of these diseases. In this regard, the solution structures of two stromelysin/ inhibitor complexes have been investigated using 1H, 13C, and 15N NMR spectroscopy. Both-inhibitors are members of a novel class of matrix metalloproteinase inhibitor that contain a thiadiazole group and that interact with stromelysin in a manner distinct from other classes of inhibitors. The inhibitors coordinate the catalytic zinc atom through their exocyclic sulfur atom, with the remainder of the ligand extending into the S1-S3 side of the active site. The binding of inhibitor containing a protonated or fluorinated aromatic ring was investigated using 1H and 19F NMR spectroscopy. The fluorinated ring was found to have a reduced ring-flip rate compared to the protonated version. A strong, coplanar interaction between the fluorinated ring of the inhibitor and the aromatic ring of Tyr155 is proposed to account for the reduced ring-flip rate and for the increase in binding affinity observed for the fluorinated inhibitor compared to the protonated inhibitor. Binding interactions observed for the thiadiazole class of ligands have implications for the design of matrix metalloproteinase inhibitors.

Binding Sites↗
PubMed1996

Biosynthesis of marcfortine A.

This report describes the results of a biosynthesis study of marcfortine A (MA). We report here that MA is derived from methionine, tryptophan, lysine and two isoprene units, the latter two being derived from acetic acid. From the 13C enrichment pattern of the pipecolic acid moeity we further conclude that this unit is derived from lysine via alpha-ketoglutarate. Therefore, we have accounted for the biogenesis of every carbon atom of MA and established the biosynthetic pathway for the pipecolic acid moiety of MA.

Acetates↗
PubMed1996

Microbiological N-deoxygenation and C-oxygenation of pioglitazone-N-oxide in a single fermentation.

Oxygenation of pioglitazone-N-oxide by a microorganism isolated from soil was accompanied by N-deoxygenation to produce the pioglitazone metabolites 5-[4-[2-[5-(1-hydroxyethyl)-2-pyridyl]ethoxy]benzyl]-2,4-thiazolidinedio ne and 5-[4-[2-(5-acetyl-2-pyridyl)ethoxy]benzyl]2,4-thiazolidinedione. The oxygenating/deoxygenating organism has been characterized as Streptomyces hygroscopicus strain 02179 (UC 11099). The culture has been deposited with Agricultural Research Service, USDA, with accession number NRRL 18975.

Fermentation↗
PubMed1992

Anthelmintic activity of dioxapyrrolomycin.

Dioxapyrrolomycin, pyrrolomycin C, pyrrolomycin D, and piericidin C2 produced by UC 11065 were evaluated as anthelmintics. Assays used to examine these compounds included effects on the free-living nematode Caenorhabditis elegans, ability to clear target nematodes (Haemonchus contortus and Trichostrongylus colubriformis) from jirds, and clearance of Haemonchus contortus from lambs. A crude extract of UC 11065 containing dioxapyrrolomycin, pyrrolomycin C, pyrrolomycin D, and piericidin C2 was active against C. elegans and against H. contortus in the jird. Purified and/or synthetic samples of dioxapyrrolomycin, pyrrolomycin C, pyrrolomycin D, and piericidin C2 were tested in the jird model; only dioxapyrrolomycin exhibited appreciable activity against H. contortus (greater than or equal to 90.9% clearance at 0.33 mg/jird), while none of the compounds showed appreciable activity against T. colubriformis. Dioxapyrrolomycin cleared 99.9% of H. contortus from lambs at 12.5 mg/kg. An in vitro migration study using susceptible and closantel-resistant H. contortus showed there is cross-resistance between dioxapyrrolomycin and closantel. Dioxapyrrolomycin appears to be a narrow-spectrum anthelmintic which works through a closantel-like mode-of-action.

Actinomycetales↗
PubMed1991

Microbial O-carbamylation of novobiocin.

Novobiocin was inactivated by Streptomyces niveus US 2094 in fermentation. The inactivation product was isolated and characterized by NMR and MS as 2"-O-carbamylnovobiocin. The MICs of novobiocin and 2"-O-carbamylnovobiocin were determined for S. niveus strains.

Chromatography↗
PubMed1991

Purification of lincosaminide O-nucleotidyltransferase from Streptomyces coelicolor Müller.

An enzyme (lincosaminide O-nucleotidyltransferase) that catalyzes 3-(5'-ribonucleotidylation) of pirlimycin and several other lincosaminide antibodies has been purified approximately 35-fold from cell-free extracts of Streptomyces coelicolor Müller NRRL 3532 (UC 5240). The crude enzyme was prepared using lysozyme and was treated with MnCl2 and (NH4)2SO4. Final purification was achieved by anion exchange chromatography. The pirlimycin reaction product was verified as being pirlimycin-3-(5'-adenylate) by NMR spectroscopy and MS. As a result of purification, this lincosaminide nucleotidylating and inactivating enzyme was separated from the macrolide phosphorylating enzyme also present in the cell-free extract.

Clindamycin↗
PubMed1990

The effect of neutral resins on the fermentation production of rubradirin.

Rubradirin is an antibiotic of complex chemical structure which is active vs. methicillin resistant staphylococci. Its development has been limited due to inadequate production yields. The incorporation of neutral resins into fermentations of Streptomyces achromogenes v. rubradiris, UC 8051 resulted in the enhanced production of rubradirin. Resins HP-20, HP-21, XAD-2, XAD-7 and XAD-16 were employed in flask and tank fermentations. The incorporation of these resins promoted 2- to 4-fold enhancements of the rubradirin activity produced in flask fermentations, and the incorporation of XAD-16 and HP-21 into tank fermentations promoted production titer increases greater than 5 fold.

Anti-Bacterial Agents↗
PubMed1990

Effects of trospectomycin on serum sensitivity of Escherichia coli UC 9451.

Trospectomycin sulfate, a chemically synthesized analog of spectinomycin, exhibits a broad range of activity against both aerobes and anaerobes, including the etiological agents of sexually transmitted diseases. Its activity in vitro against Escherichia coli is considered only moderate. At subinhibitory levels, however, trospectomycin induced changes in a pathogenic strain of E. coli, UC 9451, which significantly increased its sensitivity to serum lysis. This strain of E. coli shows high-level resistance to serum in vitro, typically growing twofold within a 45-min incubation period. Following exposure to one-fifth the MIC of trospectomycin, greater than 99% of the bacteria were killed in 25% serum within 15 min. Surviving bacteria were static in this level of serum for over 3 h. Killing was due to lysis mediated by both the classical and alternative complement pathways. The bacteria exposed to trospectomycin were enlarged in both diameter and length, but they still grew at rates comparable to those of untreated bacteria. No other visible morphological changes could be directly related to the increase in serum sensitivity. The profile of outer membrane proteins obtained by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was identical for trospectomycin-treated or untreated bacteria. However, the relative proportion of four major outer membrane proteins varied considerably.

Anti-Bacterial Agents↗
PubMed1989

Microbial glycosylation of erythromycin A.

Erythromycin A (compound 1) was inactivated by Streptomyces vendargensis ATCC 25507 in fermentation. The inactivation product was isolated and characterized by nuclear magnetic resonance and mass spectroscopy as 2'-(O-[beta-D-glucopyranosyl])erythromycin A (compound 2). The MICs of compounds 1 and 2 were determined. Compound 2 lacked antibiotic activity when tested against several gram-positive pathogens, as well as S. vendargensis.

Chromatography, Thin Layer↗
PubMed1988

Augmentation of serum bactericidal activity by paldimycin.

At concentrations below the MIC, paldimycin induced changes in Staphylococcus aureus 502A (UC 9116, ATCC 28417) which increased its sensitivity to serum. The enhanced sensitivity to serum was concentration dependent with the maximal sensitivity found when bacteria were grown in approximately 1/10 MIC of paldimycin. Within an 1-hour incubation, S. aureus 502A typically grew 1.5-2-fold in serum. Following exposure to paldimycin, however, approximately 30-50% of the bacteria were killed in serum. The paldimycin treated bacteria were not more susceptible to phagocytosis and killing by polymorphonuclear leukocytes. At the concentrations utilized, the Staphylococci were enlarged and had thickened cell walls. The organisms were still viable and replicating, but irregularities in cell division were observed in transmission electron micrographs.

Acetylcysteine↗