Search PubMed⌕ Search

Biomedical subjects

V K Mishra

Publications and source records attributed to V K Mishra.

At least 37 records · Page 2Linked to original sources
PubMed1995

Molecular basis for prokaryotic specificity of magainin-induced lysis.

Magainins and mastoparans are examples of peptide antibiotics and peptide venoms, respectively. They have been grouped together as class L amphipathic helixes [Segrest, J.P., et al. (1990) Proteins 8, 103-117] because of similarities in the distribution of Lys residues along the polar face of the helix. Class L venoms lyse both eukaryotic and prokaryotic cells whereas class L antibiotics specifically lyse bacteria. The structural basis for the specificity of class L antibiotics is not well understood. Sequence analysis showed that class L antibiotics have a Glu residue on the nonpolar face of the amphipathic helix; this is absent from class L venoms. We synthesized three model class L peptides with or without Glu on the nonpolar face: 18LMG (LGSIWKFIKAFVGGIKKF), [E14]18LMG and [G5,E14]18LMG. Hemolysis, bacteriolysis, and bacteriostasis studies using these peptides showed that the specificity of lysis is due to both the presence of a Glu residue on the nonpolar face of the helix and the bulk of the nonpolar face. Studies using large unilamellar phospholipid vesicles showed that the inclusion of cholesterol greatly inhibited leakage by the two Glu-containing peptides. These results cannot be attributed to changes in the phase behavior of the lipids caused by the inclusion of cholesterol or to differences in the secondary structure of the peptides. These results suggest that eukaryotic cells are resistant to lysis by magainins because of peptide-cholesterol interactions in their membranes that inhibit the formation of peptide structures capable of lysis, perhaps by hydrogen bonding between Glu and cholesterol. Bacterial membranes, lacking cholesterol, are susceptible to lysis by magainins.

Amino Acid Sequence↗
PubMed1995

Effect of the arrangement of tandem repeating units of class A amphipathic alpha-helixes on lipid interaction.

Exchangeable apolipoproteins possess tandem repeating units of class A amphipathic helical segments and many of them are linked together by proline residues. To understand the optimal arrangement of the amphipathic helixes for lipid association, we have studied the interactions of three model class A amphipathic helical peptides with lipids. The three peptides are: 37pA, a dimer of 18A (DWLKAFYDKVAEKLKEAF) linked together by a Pro (18A-Pro-18A); 37aA, a dimer of 18A linked together by an Ala (18A-Ala-18A); and 36A, a dimer of 18A without any linker residue (18A-18A). Circular dichroism (CD) spectra showed that the peptides are predominantly alpha-helical in aqueous and lipid environments. Temperature dependent CD studies indicated that in buffer helix stability decreases in the order 36A > 37aA > 37pA; however, in the presence of dimyristoyl phosphatidylcholine (DMPC), the above order is reversed. The retention times of the peptides on a C18 reversed-phase high performance liquid chromatography column decreased in the order 36A > 37aA > 37pA, consistent with the lengths of the nonpolar faces of the alpha-helixes being in the same order; the retention time of the parent 18A was shorter than 37pA. While 37pA adsorbed to egg phosphatidylcholine monolayers most strongly, the degree and rate of association of 36A were significantly lower. Differential scanning calorimetry indicated that, while 37pA was most effective in reducing the enthalpy of the gel to liquid-crystalline phase transition of DMPC multilamellar vesicles, 36A was least effective; 36A was even less effective than 18A. Fluorescence quenching experiments with iodide and acrylamide indicated that, in the presence of DMPC, Trp residues in 36A are most exposed to the quenchers while in 37pA they are least exposed. In the presence of DMPC, shielding of Trp in 18A from the quenchers was more than that observed with Trp residues in 36A. The results of this study suggest that the arrangement of tandem repeating amphipathic helical units which results in the formation of a class A amphipathic helix with a nonpolar face longer than five or six turns reduces the ability of the helix to associate with phospholipid.

Amino Acid Sequence↗
PubMed1995

Functional bladder neck obstruction: a rare cause of renal failure.

We studied 7 men and 6 women (average age 33 years) who presented in renal failure with obstructive voiding symptoms or retention. Of these patients 11 had a dilated upper tract and 2 had shrunken kidneys. Mean serum creatinine at presentation was 7.0 mg./dl. No abnormality was noted on cysto-panendoscopy, retrograde urethrography and voiding cystourethrography. The patients were initially treated with clean intermittent self-catheterization following 7 to 10 days of indwelling catheterization. The majority of patients had low pressure and low flow rate at initial presentation but high end filling pressure (mean 35.3 cm. water), high voiding pressure (mean 118.9 cm. water), high opening pressure (mean 95.3 cm. water) and low peak flow (mean 5.7 ml. per second) on video pressure flow electromyography. The external sphincter was relaxed during voiding but the bladder neck opened intermittently or inadequately. No proper funneling of the bladder neck was seen. Thus, functional bladder neck obstruction was considered to be responsible for obstructive voiding in these patients. Of the patients 3 void to completion with the help of alpha blockers alone, 5 underwent bladder neck incision and are voiding well, and 5 were practicing clean intermittent self-catheterization at last followup. Serum creatinine returned to near normal in 10 patients. End stage renal failure persisted in 2 patients, 1 of whom underwent renal transplantation and is voiding well but the other died without having undergone renal replacement therapy. In the remaining patient serum creatinine was stable at 3.2 mg./dl. Mean serum creatinine at 6 months of followup was 2.33 mg.%. Bladder neck obstruction is a rare cause of renal failure which can be corrected if treated appropriately.

Adult↗
PubMed1994

Interactions of synthetic peptide analogs of the class A amphipathic helix with lipids. Evidence for the snorkel hypothesis.

Class A amphipathic helixes present in exchangeable plasma apolipoproteins are characterized by the location of positively charged amino acid residues at the non-polar-polar interface and negatively charged amino acid residues at the center of the polar face. The objectives of the present study were: (i) to investigate the role of hydrocarbon side chain length of the interfacial positively charged amino acid residues in the lipid affinity of class A amphipathic helixes, and (ii) to investigate the importance of the nature of interfacial charge in the lipid affinity of class A amphipathic helixes. Toward this end, lipid interactions of the following two analogs of the class A amphipathic helix, Ac-18A-NH2 (acetyl-Asp-Trp-Leu-Lys-Ala-Phe-Tyr- Asp-Lys-Val-Ala-Glu-Lys-Leu-Lys-Glu-Ala-Phe-NH2), and Ac-18A(Lys > Haa)-NH2 (acetyl-Asp-Trp-Leu-Haa-Ala-Phe-Tyr-Asp-Haa-Val-Ala-Glu-Haa-Leu-Haa-Glu- Ala-Phe-NH2) (Haa = homoaminoalanine), were studied. The side chain of Haa has two CH2 groups less than that of lysine. The lipid affinities of these two peptide analogs were compared with that of Ac-18R-NH2, an analog of Ac-18A-NH2 with positions of the charged amino acid residues reversed. The techniques used in these studies were circular dichroism, fluorescence spectroscopy, right-angle light scattering measurements, and differential scanning calorimetry. The results of these studies indicated the following rank order of lipid affinity: Ac-18A-NH2 > Ac-18A(Lys > Haa)-NH2 > Ac-18R-NH2. These results are in agreement with the "snorkel" model proposed earlier to explain the higher lipid affinity of class A amphipathic helixes (Segrest, J. P., Loof, H. D., Dohlman, J. G., Brouillette, C. G., and Anantharamaiah, G. M. (1990) Proteins Struct. Funct. Genetics 8, 103-117). In addition, it was observed from the differential scanning calorimetry studies that Ac-18A-NH2 and Ac-18A(Lys > Haa)-NH2 interact more strongly than Ac-18R-NH2 with negatively charged dimyristoyl phosphatidylglycerol. The weaker interaction of Ac-18R-NH2 with dimyristoyl phosphatidylglycerol is suggested to be due to electrostatic repulsion between the negatively charged lipid and the interfacial negative charges of the peptide.

Amino Acid Sequence↗
PubMed1994

Should upper ureteral calculi be manipulated before extracorporeal shock wave lithotripsy? A prospective controlled trial.

Whether all upper ureteral stones must be manipulated before extracorporeal shock wave lithotripsy (ESWL*) is an ongoing controversy. In a prospective trial, symptomatic patients with solitary upper ureteral calculi less than 1 year in duration were alternated between ESWL in situ and pre-ESWL stone manipulation. Pretreatment excretory urograms were assessed for stone size and degree of proximal hydroureteronephrosis, which was graded from zero (no dilatation) to 3 (severe dilatation). Stone manipulation was done with the patient under intravenous sedation and local anesthesia. A total of 4,000 shock waves was given in a single session using Siemens Lithostar Plus, and treatment was repeated on days 4, 15 and 30 if required. The patients were evaluated 3 months after onset of therapy with excretory urogram and urine culture. Seventy patients qualified for the study (group 1-35 in situ and group 2-35 stone manipulation) and were comparable in relation to age and sex, stone size and degree of hydronephrosis. There was no significant difference between the 2 groups regarding the number of sessions (group 1-1.86 +/- 1.2 and group 2-2.03 +/- 1.2) or shock wave requirement (group 1-5,705.8 +/- 3,536.9 and group 2-5,549.1 +/- 3,837.2) for stone fragmentation. The degree of proximal dilatation did not contribute significantly towards the outcome (F ratio 0.675, p = 0.57). A total of 30 patients (85.5%) in group 1 had a satisfactory outcome at 3 months, while 3 (8.5%) had significant residual calculi and 2 stones could not be fragmented. Of the manipulated stones 33 (94%) were successfully cleared, while 2 patients required auxiliary procedures. Ureteroscopy was required in 1 patient for upward migration of the stent. Morbidity in both groups was comparable. We conclude that upper ureteral stones should be treated in situ to avoid the morbidity of manipulation.

Adult↗
PubMed1994

apoB-100 has a pentapartite structure composed of three amphipathic alpha-helical domains alternating with two amphipathic beta-strand domains. Detection by the computer program LOCATE.

Due to the great length of apolipoprotein (apo) B-100, the localization of lipid-associating domains in this protein has been difficult. To address this question, we developed a computer program called Locate that searches amino acid sequences to identify potential amphipathic alpha-helixes and beta-strands by using sets of rules for helix and strand termination. A series of model chimeric protein test datasets were created by tandem linking of amino acid sequences of multiple proteins containing four different secondary structural motifs: motif A (exchangeable plasma apolipoproteins); motif G (globular alpha-helical proteins); motif C (coiled-coil alpha-helical proteins); and motif B (beta pleated-sheet proteins). These four test datasets, as well as randomly scrambled sequences of each dataset, were analyzed by Locate using increasingly stringent parameters. Using intermediately stringent parameters under which significant numbers of amphipathic helixes were found only in the unscrambled motif A, two dense clusters of putative lipid-associating amphipathic helixes were located precisely in the middle and at the C-terminal end of apoB-100 (a sparse cluster of class G* helixes is located at the N-terminus). The dense clusters are located between residues 2103 through 2560 and 4061 through 4338 and have densities of 2.4 and 2.2 amphipathic helixes per 100 residues, respectively; under these conditions, motif A has a density of 1.4 amphipathic helixes per 100 residues. These two domains correspond closely to the two major apoB-100 lipid-associated domains at residues 2100 through 2700 and 4100 through 4500 using the principle of releasability of tryptic peptides from trypsin-treated intact low-density lipoprotein. The classes of amphipathic helixes identified within these two putative lipid-associating domains are considerably more diverse than those found in the exchangeable plasma apolipoproteins. Interestingly, apoB-48 terminates at the N-terminal edge of the middle cluster. By using a similar strategy for analysis of amphipathic beta-strands, we discovered that the two gap regions between the three amphipathic helix clusters are highly enriched in putative amphipathic beta-strands, while the three amphipathic helical domains are essentially devoid of this putative lipid-associating motif. We propose, therefore, that apoB-100 has a pentapartite structure, NH2-alpha 1-beta 1-alpha 2-beta 2-alpha 3-COOH, with alpha 1 representing a globular domain.

Algorithms↗
PubMed1994

99m Tc DTPA scan as a diagnostic marker of acute rejection in renal transplantation.

Forty renal transplant recipients were subjected to 99m Technicium Diethylene triamine pentacetic acid (DTPA)scans at regular intervals & whenever there was suspection of rejection. Serial scans of a group of 15 recipients from immediate post transplant period till withdrawal of cyclosporine were analysed separately & the results compared to with single scan analysis. The sensitivity & specificity of DTPA scan in the absence of acute tubular necrosis (ATN) was 94.1% & 87.5%, while the positive & negative predictive values were 88.8 & 93.3% respectively. Sensitivity & specificity of serial scan analysis (88.8% and 75%) in early post transplant period was higher than that of interpretation of single scan (75% & 66%). Serial scan changes predated clinical rejection during cyclosporine withdrawal period. We conclude that DTPA scan is both a sensitive & specific non-invasive diagnostic marker of acute rejection in absence of ATN & serial scans during early post transplant period & at the time of cyclosporine withdrawal are helpful in detecting the rejection accurately & at the earliest.

Adult↗
PubMed1993

Reciprocal effects of apolipoprotein and lytic peptide analogs on membranes. Cross-sectional molecular shapes of amphipathic alpha helixes control membrane stability.

Apolipoprotein (class A) amphipathic helixes are postulated to act as detergents by virtue of their cross-section being wedge-shaped. Using computer analysis of naturally occurring class A and lytic (class L) amphipathic helixes, we designed two archetypical model peptides. Analogs of these two peptides, incorporating substitutions or modifications of interfacial or basic residues, had the following effects. Class A peptides stabilized bilayer structure, reduced leakage from large unilamellar vesicles and erythrocytes, and inhibited lysis induced by class L peptides. Class L peptides destabilized bilayer structure in model membranes and increased binding of class A peptides to erythrocytes. The ability of class L analogs to lyse membranes and induce inverted lipid phases was reduced by either decreasing the bulk of an interfacial residue, increasing the angle subtended by the polar face, or increasing the bulk of the basic residues. The ability of the class A analog to stabilize bilayer structure and inhibit erythrocyte lysis by class L peptides was enhanced by methylating the Lys residues. These results can be explained by a model that we term the reciprocal wedge hypothesis. By analogy to the reciprocal effects of phospholipid shapes on membrane structure, we propose that the wedge shape of class A helixes stabilizes membrane bilayers, whereas the inverted wedge shape of class L helixes destabilizes membrane bilayers, and, thus, one class will neutralize the effect of the other class on membranes.

Amino Acid Sequence↗
PubMed1993

Testicular toxicity of thiram in rat: morphological and biochemical evaluations.

Tetramethylthiuram disulphide (Thiram), a dithiocarbamate fungicide was administered to male rats through gavage at dose 5, 10 and 25 mg/kg/day for 30, 60 and 90 days. A significant increase in relative testes weight (GSI) was observed at dose (25 mg/kg/d) after 90 days exposure, associated with mild pathomorphological changes. This included degeneration of seminiferous tubules with sloughing of germinal cells and accumulation of edematous fluid in tubular lumen. The activity of testicular enzymes such as lactate dehydrogenase (LDH), alkaline phosphatase (ALP), glucose 6-phosphate dehydrogenase (G-6-PDH) were increased significantly in dose dependent manner. The activities of succinate dehydrogenase (SDH) and acid phosphatase (ACP) decreased significantly while serum cholesterol and testicular free sialic acid were increased at all dose levels. The alterations in the activity of testicular enzymes associated with marked increase in GSI and pathomorphological changes suggest that thiram induces testicular dysfunctions in rats.

Animals↗
PubMed1993

Significance of serum immunoglobulin levels in conglomeration with trans-sternal phlebography in the phlebography management of breast cancer.

The study was conducted on 63 patients with breast lump and twenty normal healthy females. In benign breast disease, a significant (P < 0.001) rise in serum IgA, significant (P < 0.001) decrease in IgG and no change in IgM levels was seen before operation. A significant decrease (P < 0.001) in serum IgA and significant increase in IgG and IgM was observed post operatively. In carcinoma breast, a significant (P < 0.001) elevation in IgA, IgG and IgM levels was found pre-operatively with a concomitant decrease in serum IgA and IgG and increase in serum IgM levels, post operatively. Trans-sternal phlebography (TSP) carried out with 95.23% success has revealed significant (P < 0.001) change in the staging of carcinoma breast. The increased levels of serum immunoglobulins associated with the patients of carcinoma breast with metastasis has led to conclude that these levels, if punctuated with TSP findings can lead to better assessment of the staging of carcinoma breast and thereby its management.

Biomarkers, Tumor↗
PubMed1992

Routine radio-imaging in filarial chyluria--is it necessary in developing countries?

Chyluria is a common presentation of filariasis in developing countries and mostly affects economically poor classes. The judicious use of investigations is mandatory to keep the cost of treatment acceptable. A modified plan is suggested for managing such patients without using retrograde pyelography or lymphangiography. A total of 54 patients were studied at 2 separate institutions, with retrograde pyelography being carried out at one institution but not at the other. All patients were treated primarily by the instillation of 1% silver nitrate, with an initial success rate of 70%. Eight patients who failed to respond underwent pyelolymphatic disconnection, the success rate being 88%. The omission of retrograde pyelography and lymphangiography had no effect on either treatment or outcome. The value of these radio-imaging techniques is discussed.

Adult↗