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Biomedical subjects

S Joshi

Publications and source records attributed to S Joshi.

At least 199 records · Page 11Linked to original sources
PubMed1988

Cloning of the genes of the chitin utilization regulon of Serratia liquefaciens.

The set of genes that determine the expression of the enzymes involved in chitin degradation by Serratia liquefaciens was cloned. The role of each gene was investigated, and for the first time regulatory genes were identified in this system. The chiA and chiB genes coded for separate chitinase activities. The chiC region coded for a chitobiase activity, but it was not formally separated from chiB. Transposon mutagenesis and deletion analysis identified a region, chiD, whose absence led to higher expression of chiA, chiB, and chiC. chiD may therefore be a gene that codes for a repressor. Loss of function of another adjacent region, chiE, prevented induction unless a chiE+ strain was a near neighbor, suggesting that this gene may code for a protein that is involved in the synthesis of the inducer. chiB, chiC, chiD, and chiE are closely linked, while chiA is in a separate location on the chromosome.

Acetylglucosaminidase↗
PubMed1988

Renal ammoniagenic response to chronic acid loading: role of glucocorticoids.

Adrenalectomized (ADX) animals exhibit a blunted renal response to chronic acid loading. To determine whether this response truly reflects impaired renal ammoniagenesis from glutamine, urinary ammonium excretion was compared with acid intake in ADX, intact, and ADX rats supplemented with either a low dose (4 micrograms.100 g-1.day-1) or a high dose (40 micrograms.100 g-1.day-1) of triamcinolone. ADX rats consumed similar amounts of acid as did intact controls yet excreted only 37% of the load as ammonium; in contrast intact controls returned 86% and triamcinolone-supplemented animals returned 98 and 88% for low and high doses, respectively. Nor could the reduced ammonium excretion be attributed to increased renal venous release, since total ammonia production, the sum of renal venous and urine ammonium, was reduced to 49% of the intact controls; low- and high-dose triamcinolone restored and markedly increased the production rate. Underlying the impaired ammonia production rate in ADX rats was a reduced rate of glutamine extraction, 350 +/- 49 vs. 896 +/- 102 and 1,260 +/- 247 and 1,448 +/- 112 nmol.min-1.100 g-1 for intact and low and high doses, respectively. Unlike intact acidotic and glucocorticoid-supplemented ADX acidotic rats, glutamine extraction was disassociated from the delivered glutamine load consonant with the role of glucocorticoid in coupling cellular glutamine transport to its metabolic utilization.

Acid-Base Equilibrium↗
PubMed1988

Assessing supplementary feeding programmes in selected Balwadies.

The aim of the study was to investigate whether feeding programmes achieve their goal of improving nutritional status of target groups when such groups live in environmental conditions that constantly pose risk of infections and infestations. Two major areas from Pune city were selected: one comprised mostly slum dwellers, living in poor environmental conditions (LSE) and the other with better-off families living in good environmental conditions (MSE). From each of the two areas two Balwadies (kindergartens) were selected: one was implementing a feeding programme (LSE(F) and MSE(F)) and the other was not (LSE(NF) and MSE(NF]. A total of 273 newly admitted pre-school children from these Balwadies were selected and anthropometric measurements were made three times during the study period of 7 months. Food items distributed in the school were weighted and analysed for energy and protein values. Assessment of nutritional status by the Gomez classification showed that in both areas, moderately and severely malnourished children receiving the supplement showed significant improvement (P less than 0.05) as compared to those not receiving the supplement, but the extent of improvement differed. The impact of the food supplement in LSE was less than that in MSE suggesting that poor environmental conditions were a major interfering factor and cannot be overlooked.

Anthropometry↗
PubMed1987

Hybrid molecules containing the A-domain of insulin-like growth factor-I and the B-chain of insulin have increased mitogenic activity relative to insulin.

Two synthetic insulin-like compounds consisting of the B-chain of insulin linked via disulfide bonds to A chains corresponding to the A-domain or the A- and D-domains of insulin-like growth factor I (IGF-I) have been evaluated for mitogenic activity and for binding to IGF receptors and IGF carrier proteins. Both compounds are 3- to 5-fold more potent mitogens than insulin, and have a comparably increased affinity for the type I IGF receptor that mediates these mitogenic effects in chick embryo fibroblasts. Neither compound interacts with IGF carrier proteins. These results indicate that the A-domain of IGF-I is importantly involved in its growth-promoting properties.

Animals↗
PubMed1987

Renal brush border glutamine transport: comparison between in situ and isolate membrane vesicle uptake.

Glutamine uptake by renal cortical brush-border vesicles was compared to transport expressed by the functioning isolated kidney. Comparisons were made with regard to sodium dependency and the adaptive increase induced by chronic metabolic acidosis in the rat. The results show an absolute dependency upon a sodium gradient; sodium-independent glutamine uptake has no counterpart in situ. In addition, acidosis-induced adaptive increase in vesicle glutamine uptake has no counterpart in situ. Rather, the apparent adaptation reflects extravesicular gamma-glutamyltransferase-mediated conversion to glutamate and subsequent accumulation; acidosis-induced adaptation of this enzyme largely explains the apparent adaptation in glutamine uptake. Consequently the role of membrane transport in glutamine flux regulation can be assessed providing metabolic conversion is controlled.

Acidosis↗
PubMed1987

Specificity of fungal lipase in hydrolytic cleavage of oil.

Lipase isolated from the fungus Fusarium oxysporum f. sp. lini has shown specificity to saturated acids. In vitro hydrolytic splitting of cotton seed, ground-nut and fungal (F. oxysporum) oil substrates by the enzyme preparation from this fungus, revealed preferential fatty acid specificity. The enzyme attacked the triglyceride molecule, liberating 90-97% of saturated fatty acids, irrespective of length of the carbon chain.

Arachis↗
PubMed1986

Topology and function of "stalk" proteins in the bovine mitochondrial H+-ATPase.

Proton translocating ATPases comprise a hydrophilic sector F1, a membrane sector F0, and, in the case of bovine mitochondria, a connecting "stalk" which is believed to contain the oligomycin sensitivity-conferring protein (OSCP) and coupling factor 6 (F6). The present study was undertaken to verify the accessibility of F6 and OSCP to trypsin and to examine the functional consequences of such treatment. Our data show that F1 binds equally to trypsin-treated F0 and untreated F0, but the former complexes exhibit cold lability and only partial sensitivity to oligomycin. Furthermore, these complexes fail to exhibit ATP-driven proton translocation or ATP-32Pi exchange activity. Trypsinization of F0 does not, however, inhibit passive proton conductance through the membrane sector but actually enhances it. Immunological data indicate extensive degradation of OSCP under conditions where F6 proteolysis is insignificant. Intact H+-ATPase complexes are relatively resistant to both the structural and functional effects of trypsin. We conclude that OSCP is predominantly an extrinsic protein which is shielded by F1 in the native membrane. F6 may also be an extrinsic protein but is shielded from trypsinization by OSCP and/or other F0 polypeptides. The exposed, trypsin-sensitive segments of OSCP are not required for passive proton conductance through F0 but may be required for ATP-driven reactions. We propose that bovine mitochondrial OSCP is a functional analogue of subunit b in the Escherichia coli H+-ATPase.

Adenosine Triphosphatases↗
PubMed1986

Regulation of interorganal glutamine flow in metabolic acidosis.

Metabolic acidosis redirects interorgan glutamine flow from hepatic utilization to renal ammoniagenesis at the expense of ureagenesis. The roles of arterial glutamine load and organ glutaminase capacity in the regulation of glutamine balance across the gut, liver, and kidneys were studied in control and chronically acidotic rats. In control rats these organs combined to remove 733 nmol glutamine X min-1 X 100 g-1 in agreement with their respective glutaminase content, gut greater than liver greater than kidneys. In chronic metabolic acidosis renal glutamine extraction alone increased to 1,158 nmol X min-1 X 100 g-1 associated with an increased glutaminase capacity. However, the total glutamine deficit across these organs rose to only 1,043 nmol glutamine consumed X min-1 X 100 g-1 as a consequence of hepatic glutamine uptake reversing to net release. This reversal was not dependent on increased hepatic glutamine synthetase capacity, but rather appears to be dependent on the combined effect of reduced portal venous glutamine load and increased ammonia load. The reduction in portal glutamine load is, in turn, a consequence of renal glutamine extraction and reduced arterial glutamine concentration in metabolic acidosis as well as maintained gut glutamine extraction. Elevating arterial glutamine concentration in metabolic acidosis has no effect on renal uptake, but enhances splanchnic bed extraction with the restoration of ureagenesis. Thus the interorgan flow of glutamine and deposition of N into either urea or ammonia appears to be dependent on arterial glutamine concentration and hence glutamine availability in chronic metabolic acidosis in the rat.

Acid-Base Equilibrium↗
PubMed1986

Mitogenic activity and receptor reactivity of hybrid molecules containing portions of the insulin-like growth factor I (IGF-I), IGF-II, and insulin molecules.

Insulin and the insulin-like growth factors IGF-I and IGF-II are thought to exert their mitogenic effects in cultured chick embryo fibroblasts and human skin fibroblasts via IGF receptors rather than via insulin receptors. These effects appear to be mediated by the type I subtype of IGF receptor, which is structurally similar to the insulin receptor and exhibits significant cross-reactivity with insulin. As a first step in our long-range goal of defining those features of the IGF-I and IGF-II molecules that confer enhanced mitogenic activity and reactivity with these mitogenic type I IGF receptors, we have prepared two hybrid insulin-IGF molecules and examined their mitogenic and binding activities: (1) A27-insulin, containing an elongated 27-residue A-chain (in which the 6-residue D-domain of IGF-II was added to the carboxy-terminus of the 21-residue A-chain of insulin) combined with the B-chain of insulin; and (2) A insulin-B IGF-1, containing the A-chain of insulin and the synthetic 30-residue B-domain of IGF-I. Both hybrid molecules stimulated DNA synthesis and inhibited 125I-IGF-I binding to type I IGF receptors in both chick embryo and human fibroblast cultures. A27-insulin had considerably greater mitogenic potency and binding potency than A insulin-B IGF-I. Neither hybrid molecule was more potent in these assays than insulin, indicating that the presence of D IGF-II or B IGF-I by itself was not sufficient to increase the mitogenic potency of insulin in fibroblasts. By contrast, A insulin-B IGF-I showed enhanced reactivity with an antiserum to IGF-I. A27-insulin retained significant insulin-like metabolic activity despite the presence of the D-domain of IGF-II.

Animals↗
PubMed1986

Hepatic enzymes of glutamine and ureagenesis in metabolic acidosis.

The activities of key glutamine and urea cycle enzymes were assayed in liver homogenates from control and chronically acidotic rats and compared with citrulline and urea productions by isolated mitochondria and intact liver slices, respectively. Glutamine-dependent urea and citrulline synthesis were increased significantly in isolated mitochondria and in liver slices; the activities of carbamoyl phosphate synthetase and arginase were unchanged and increased, respectively. Glutamine was not a precursor in the carbamoyl phosphate synthetase system, suggesting that the glutamine effect is an indirect one and that glutamine requires prior hydrolysis. Increased mitochondrial citrulline synthesis was associated with enhanced oxygen consumption, suggesting glutamine acts both as a nitrogen and fuel source. Hepatic phosphate-dependent glutaminase was elevated by chronic acidosis. The results indicate that the acidosis-induced reduction in ureagenesis and reversal from glutamine uptake to release observed in vivo are not reflections of corresponding changes in the hepatic enzyme content. Rather, when available, glutamine readily supports ureagenesis, suggesting a close coupling of hepatic glutaminase flux with citrulline synthesis.

Acidosis↗
PubMed1985

Structural features involved in the biological activity of insulin and the insulin-like growth factors: A27 insulin/BIGF-I.

A synthetic insulin-like compound consisting of the A-chain of insulin extended at its carboxyl terminus with the hexapeptide "D-domain" of insulin-like Growth Factor II, linked via disulfide bonds to a B-chain corresponding to the "B-domain" of insulin-like Growth Factor I, has been examined for insulin-like metabolic activity and for mitogenic activity. The synthetic material (A27 insulin/BIGF-I) is less potent than insulin in metabolic assays, and less potent than both insulin and IGF-I in mitogenic assays. It is proposed that neither the "D-domain" nor the "B-domain" of the IGFs is a major contributor to mitogenic activity. Their presence in the same molecule does not result in significant growth-promoting activity.

Adipose Tissue↗
PubMed1985

Cross-linking of bovine mitochondrial H+-ATPase by copper--o-phenanthroline. Interaction of the oligomycin-sensitivity-conferring protein with a 24-kDa protein.

The nearest neighbor relationships between the Fo subunits of bovine mitochondrial H+-ATPase were studied by using copper-o-phenanthroline, an SH-oxidizing cross-linking reagent. The cross-linked samples of purified H+-ATPase, F1-ATPase or Fo were analyzed by sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS-PAGE) and the disulfide cross-linked polypeptides were identified by enzyme-linked immunosorbent assay and immunoblot transfer using subunit specific antisera. SDS-PAGE of H+-ATPase showed several cross-links, although none involved subunits of Fo sector linked to those of F1. Both H+-ATPase and Fo showed formation of a 45-kDa product. Upon reduction, the 45-kDa component gave rise to a 21-kDa band, identified as oligomycin-sensitivity-conferring protein (OSCP), and a 24-kDa band. These two proteins thus appear to be near neighbors with their cysteine residues in close proximity with each other. Under the conditions of cross-linking, there was a concentration-dependent decrease in the Pi-ATP exchange activity of the intact H+-ATPase as well as of H+-ATPase reconstituted with copper-o-phenanthroline-treated Fo and untreated F1. The site of inhibition appeared to residue in the Fo sector. Loss of Pi-ATP exchange occurred at the same time as formation of the 45-kDa product. Our present data showing copper-o-phenanthroline-induced interactions of the 24-kDa protein with the OSCP and simultaneous inactivation of Pi-ATP exchange activity of the complex strengthen earlier suggestions [Hadikusumo, R.G., Hertzog, P.J. & Marzuki, S. (1984) Biochim. Biophys. Acta 765,258-267] that the 24-kDa protein may be a bona fide subunit of Fo.

Adenosine Triphosphatases↗
PubMed1985

Structures of shorthorn sculpin antifreeze polypeptides.

The amino acid sequences of the two major antifreeze polypeptides (AFP) from the shorthorn sculpin have been determined using an automatic protein sequencer and enzymic digestion. These two polypeptides, SS-3 and SS-8, consist of 33 and 45 amino acid residues respectively. The N-terminal methionyl residue is blocked in both the polypeptides. When aligned for maximum structural similarity these two AFP are 80% homologous, and there appears a deletion of 12 amino acid residues at the N-terminal portion of SS-3. Like the winter flounder AFP, both the sculpin AFP also contain the 11-amino-acid repeat sequences. The secondary structure of the sculpin AFP is mainly alpha-helical as deduced from circular dichroic spectral data. The helical content of SS-8 is high (73%), while that of SS-3 is moderate (about 45%). The latter exhibits a relatively weak antifreeze activity. Removal of the blocked N-terminal residue in SS-8 did not alter the helical content significantly but did reduce the antifreeze activity. Helical contents of proteolytically generated fragments of AFP are much lower, and they are devoid of activity. The alpha-helix in the SS-8 component is seen to be amphiphilic in character. The relevance of this feature to the mechanism of the antifreeze action is briefly discussed.

Amino Acid Sequence↗
PubMed1985

Synthesis of an insulin-like compound consisting of the A chain of insulin and a B chain corresponding to the B domain of human insulin-like growth factor I.

An insulin-like hybrid molecule consisting of the A chain of insulin and a B chain corresponding to the B domain of human insulin-like growth factor I (growth factor I sequence 1-30) has been synthesized essentially by the procedures developed in this laboratory for the synthesis of insulin and analogues. The hybrid competed with 125I-insulin for insulin receptors in rat liver plasma membranes and was a full agonist in stimulating incorporation of [3(-3)H]glucose into lipids in rat adipocytes. In both assays, the compound displayed ca. 2% of the potency of insulin. The compound was recognized by anti-insulin antibodies but was only ca. 0.25% as potent as insulin in this activity. The hybrid exhibited growth-promoting activity in fibroblasts, displaying 3-8% of the activity of insulin. In contrast, the compound was recognized by insulin-like growth factor carrier proteins, a property not associated with insulin. Two points of nonhomology between the B chain of insulin and the B domain of insulin-like growth factor I are considered in connection with these observations.

Adipose Tissue↗
PubMed1985

Formation of an intramolecular disulfide bond in the mitochondrial adenine nucleotide translocase.

Adenine nucleotide translocase in electron transport particles or in H+-ATPase preparation from bovine heart mitochondria is capable of forming both inter- and intramolecular disulfide bridges upon reaction with copper-o-phenanthroline. We have examined the localisation of the intramolecular disulfide bridge in the protein chain by peptide fragmentation methods. The most likely position of the disulfide bridge is between cysteine 159 and 256, but the possibility of the presence of a second disulfide bridge formed between 129 and 256 cannot be ruled out. Our experimental results support the theoretical model proposed [(1982) FEBS Lett. 144, 250-254] for the topography of the translocase and provide a more accurate description of the arrangement of some of the hydrophilic segments in the molecule.

Animals↗
PubMed1985

Monoclonal antibodies to mitochondrial coupling factor B.

Two monoclonal antibodies (MAb I and IV) have been prepared which showed high and specific reactions towards bovine heart mitochondrial coupling factor B (FB). Both have been identified as sub-type IgG1 of mouse immunoglobulins. MAb I reacts with purified and functionally active FB, alkylated or oxidized forms of FB and even with peptides formed on digestion of FB with trypsin. When used together, MAb I and IV reacted with FB in immunoblots of normal and urea treated samples of mitochondria, submitochondrial particles, ammonia-EDTA extracted particles, and H+-ATPase. Both MAbs inhibited FB-stimulated ATP-dependent reverse electron flow activity when FB was incubated with the antibody either before or after its addition to FB-deficient AE-particles. Reactivity of MAb I towards FB declined upon exposure of FB to guanidine HC1 while reactivity of MAb IV remained unaltered.

Animals↗
PubMed1985

Hybrid molecules containing the B-domain of insulin-like growth factor I are recognized by carrier proteins of the growth factor.

The insulin-like growth factors (IGFs) are polypeptides in plasma that are chemically related to insulin and have mitogenic and insulin-like activity. Unlike insulin, the IGFs circulate in plasma bound to specific high molecular weight carrier proteins that regulate their delivery to target tissues. To define the sites on the IGFs that allow them to be recognized by carrier proteins, we constructed hybrid molecules containing different portions of the insulin, IGF-I, and IGF-II molecules. The presence of the B domain of IGF-I, but not the D domain of IGF-II, enables these insulin-IGF hybrid molecules to be recognized by acid-stripped IGF carrier proteins from rat serum and other sources. By contrast, neither the BIGF-I nor DIGF-II domain is sufficient to enable binding to type II IGF receptors, despite the fact that type II receptors, like the carrier protein, specifically bind IGF-I and IGF-II but do not interact with insulin. By differentiating those sites on the IGF molecule required for binding to IGF carrier protein and receptors, the insulin-IGF hybrid molecules should help delineate the role of the carrier protein in presenting biologically active IGF to target tissues.

Animals↗