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Biomedical subjects

E Mendez

Publications and source records attributed to E Mendez.

At least 55 records · Page 3Linked to original sources
PubMed1990

Complete purification of two identical Na(+)-pump inhibitors isolated from bovine hypothalamus and hypophysis.

We have completely purified, in parallel, a low molecular weight, non-specific, non-lipidic, Na+,K(+)-ATPase inhibitory factor from bovine hypothalamic and pituitary tissues. In the final purification step we obtain, from both tissues, a single, homogeneous peak, with a maximal absorbance at 247 nm. This factor, at physiological concentrations of potassium (5-25 mM), inhibits in a dose-response manner Na+,K(+)-ATPase and displaces ouabain from its receptor at the enzyme structure. The factor isolated from both tissues is identical, being the specific activity per weight of tissue higher in hypophysis. No factor was found in cerebral cortex, used as tissue control.

Animals↗
PubMed1989

Heat-stable translational inhibitor from rabbit reticulocyte lysates.

We have purified to apparent homogeneity a heat-stable (HS) factor from the postribosomal supernatant of rabbit reticulocyte lysates [(1988) FEBS Lett. 236, 479-483]. HS inhibits translation in hemin-supplemented lysates and induces phosphorylation of the alpha-subunit of the eukaryotic initiation factor 2 as does hemin deficiency. The translational inhibition produced by addition of HS to hemin-containing reticulocyte lysates and the accompanying phosphorylation of the eIF-2 alpha subunit can be prevented or reversed by NADPH generators including glucose 6-phosphate, NADPH itself, and also by dithiols, e.g., dithiothreitol, but not by fructose 1,6-bisphosphate or by monothiols, e.g., 2-mercaptoethanol. When added to crude preparations of the proinhibitor form (proHCI) of the heme-controlled translational inhibitor (HCI), HS produces a pronounced increase of the HCI to proHCI ratio. It appeared possible that HS might be oxidized glutathione (GSSG) but this is not the case, for HS is not a substrate for highly purified glutathione reductase from rabbit erythrocytes. The spectral analysis of highly purified HS is consistent with the idea that HS could be a nucleotide derivative.

Animals↗
PubMed1989

Structural and idiotypic characterization of the L chains of human IgM autoantibodies with different specificities.

We have determined the V region amino acid sequence and/or serologic markers (kIIIb, PSL2, and PSL3) of 24 IgM monoclonal autoantibodies with specificities of anti-gamma-globulin (RF), anti-I (cold agglutinin), anti-low density lipoprotein and anti-intermediate filaments. The data emphasize the overwhelming selection of the HumKv325/VkIIIb L chain for this family of autoantibodies. The few amino acid substitutions found within the VL regions were mainly concentrated in the complementarity-determining region 1. JK and CK genes did not show the same pattern of restriction. There is a good correlation between the amino acid sequence and the presence of the kIIIb marker. The idiotypic marker PSL2 was present in 34 out of 35 kIIIb L chains analyzed (97%) and the PSL3 in 27 (80%). Moreover, the hydrophilicity and antigenic profiles of these L chains corroborate the presence of the epitopes detected by the anti-CRI. These results demonstrate a restricted selection of the Vk genes used by a family of self reacting proteins, and an unusual evolutionary conservation of the idiotypic structure that may be involved in the network regulation.

Agglutinins↗
PubMed1989

Purification and properties of five different forms of human procarboxypeptidases.

Three different procarboxypeptidases A and two different procarboxypeptidases B have been isolated for the first time, in a pure and native state, from human pancreatic extracts. These proteins were purified in one or two quick steps by anion-exchange HPLC. All these forms have been biochemically characterized. Two of the procarboxypeptidases A, the A1 and A2 forms, are obtained in a monomeric state while the other, the A3 form, is obtained as a binary complex of a procarboxypeptidase A with a proproteinase E. This complex is stable in aqueous buffers at various ionic strengths and develops carboxypeptidase A and proteinase E activities in the presence of trypsin. The A1 and A2 forms show clear differences in electrophoretic mobility in SDS/polyacrylamide gels, isoelectric point, proteolytic activation process with trypsin and susceptibility to thermal denaturation. In contrast, these properties are similar in the A1 and A3 (binary complex) forms. On the other hand, with respect to the properties listed above, the B1 and B2 forms differ from each other mainly in isoelectric point. An overall comparison of the above properties reveals the unusual character of the A2 form, midway between the other A and B forms. N-terminal extended sequence analysis carried out on these proenzymes confirm that they constitute different isologous forms.

Animals↗
PubMed1989

Naturally occurring serotype 2/subgroup II rotavirus reassortants in northern Brazil.

Nine serotype 2 human rotavirus strains were isolated in a community-based longitudinal study in Northern Brazil. Five of these strains had a 'long' RNA electrophoretic pattern and all five strains were determined to belong to subgroup II by ELISA assay, in contrast to properties common to serotype 2 human rotaviruses previously characterized. Hybridization studies of one of these unusual strains with 32P-labelled mRNAs derived from the prototype human strains Wa (serotype 1, subgroup II) and S2 (serotype 2, subgroup I) suggested that it was generated by a reassortment event in nature, in which a subgroup II, 'long' electropherotype rotavirus exchanged its serotype-specific gene and gene number 10 for the equivalent genes from a serotype 2, 'short' electropherotype virus.

Brazil↗
PubMed1989

Serum levels of protein HC and its IgA complex in malignant melanoma.

Protein HC is a low molecular weight glycoprotein that is present as free form and in complex with IgA (HC-IgA) in human body fluids. In the present report, a combined competitive and sandwich enzyme-linked immunsorbent assay (ELISA) was used to measure the levels of protein HC and its IgA complex independently in the sera of patients with malignant melanoma. The concentration of these proteins correlated with IgA levels and clinical staging. The main findings can be summarized as follows: (1) protein HC and protein HC-IgA complex levels presented the most significant elevation in melanoma stages IVA and IVB; (2) a significant positive correlation was found between protein HC and protein HC-IgA complex values and the clinical stage of the tumor; (3) protein HC-IgA complex and IgA concentrations are parameters with positive correlation both in normal controls and in patients with melanoma; (4) in normal sera, the levels of protein HC and protein HC-IgA complex are not correlated, however, they are negatively correlated in melanoma.

Adult↗
PubMed1989

Synthesis and apparent secretion of prothymosin alpha by different subpopulations of calf and rat thymocytes.

Prothymosin alpha (ProT alpha), a polypeptide containing the sequence of thymosin alpha 1 (T alpha 1) at its NH2-terminus, has been isolated from calf thymocytes in a concentration in the order of that found in the whole thymic gland. As deduced from the analysis of their tryptic peptides, calf ProT alpha differs from the rat polypeptide at least in an alanine residue replacing valine at position 92. Thymocytes cultured in a radioactive medium exhibit an important secretory activity, ProT alpha being one of the products synthesized and exported to the culture medium. Large and small thymocyte subpopulations from calf and rat differ in their capacity to synthesize ProT alpha. The polypeptide is produced in a major concentration by large thymocytes. However, all the calf and rat thymocyte subpopulations show a similar capacity to secrete ProT alpha, the amount of the newly synthesized polypeptide recovered from cell culture supernatants being 80-90% of that found in thymocyte extracts.

Animals↗
PubMed1988

Purification of a novel heat-stable translational inhibitor from rabbit reticulocyte lysates.

We have purified to apparent homogeneity a novel heat-stable (HS) factor from postribosomal supernatants of rabbit reticulocyte lysates by heating for 10 min at 80 degrees C, fractionation on Sephadex, anion-exchange chromatography on QMA Accell, and gel filtration HPLC. The apparent molecular mass of HS is 500-1000 Da on the basis of its behaviour on gel filtration. Like a factor from bovine heart [(1982) Proc. Natl. Acad. Sci. USA 79, 3134-3137], the reticulocyte HS inhibits translation in hemin-supplemented lysates with biphasic kinetics similar to hemin deficiency and promotes phosphorylation of the alpha-subunit of the eukaryotic initiation factor eIF-2. It is active at nanomolar concentrations. Reticulocyte HS appears to be neither a peptide nor an oligonucleotide since HS activity was insensitive to proteolytic or nucleolytic digestion.

Animals↗
PubMed1988

Specific phosphorylation of the beta subunit of eIF-2 factor from brain by three different protein kinases.

The eukaryotic initiation factor 2 (eIF-2) from calf brain has been purified to homogeneity and free of endogenous kinase activity. Phosphorylation of eIF-2 factor has been examined with four different protein kinases. Casein kinase II, calcium/phospholipid-dependent protein kinase and cyclic AMP-dependent protein kinase from brain, phosphorylate the beta subunit of eIF-2, whilst hemin-controlled inhibitor phosphorylate the alpha subunit of the factor. According to the peptide maps obtained, the phosphorylation sites of the factor by the three beta kinases are specific and distinct. These data suggest a different regulation for the beta subunit through this modification.

Animals↗
PubMed1988

Improvement in HPLC fractionation of thyroxine-containing thyroglobulin tryptic peptides by prior Accell ion-exchange column chromatography.

The isolation and purification of the peptides containing the hormonogenic tyrosyl residues in thyroglobulin is of great interest to the understanding of structure-function relationships in this iodoprotein. This is usually performed in reduced alkylated selectively hydrolyzed thyroglobulin by subsequent HPLC fractionation. However, the main difficulty encountered when trying to isolate these peptides is their disproportion with respect to the total number of possible peptides (14 vs a total of 508). Several HPLC runs with different mobile phases are necessary and each run is accompanied with significant losses of the peptides to be purified. In an attempt to improve the separation of the T4-containing peptides in thyroglobulin tryptic digests from the much more abundant iodotyrosine-containing ones, which are present as contaminants, we have used a very simple and fast step prior to the HPLC fractionation as it is a self-packed ion-exchange column chromatography. This preliminary step results in an improvement in the separation of these peptides and leads to a relative enrichment of the hormonogenic peptides falling in different zones of the HPLC chromatogram, which facilitates their subsequent separation and purification by HPLC.

Acetonitriles↗
PubMed1986

New alpha-amylase and trypsin inhibitors among the CM-proteins of barley (Hordeum vulgare).

Barley CM-proteins are a group of at least five salt-soluble components (CMa-e) that can be selectively extracted from endosperm with chloroform/methanol mixtures. N-terminal sequences of proteins CMa, CMb and CMc have been determined and found to be homologous to those previously determined for CMd and CMe, an observation which confirms that their structural genes are members of a dispersed multi-gene family. The purified CM-proteins were tested against trypsin and against alpha-amylases from saliva, pancreas, Aspergillus oryzae, Tenebrio molitor and barley. Besides CMe, which was known to be a trypsin inhibitor, CMc also showed antitrypsin activity, whereas CMa was specifically active against the alpha-amylase from T. molitor and no inhibitory activity was found for proteins CMb and CMd. The evolutionary implications of these findings are discussed.

Amino Acid Sequence↗
PubMed1985

Differential effects of high-lysine mutations on the accumulation of individual members of a group of proteins encoded by a disperse multigene family in the endosperm of barley (Hordeum vulgare L.).

The CM proteins are a group of major salt-soluble endosperm proteins encoded by a disperse multigene family. The effects of high-lysine mutations on the net accumulation in barley endosperm of three members of this group (CMa, CMb, and CMe) have been investigated. Genes CMa, CMb and CMe are located in chromosomes 1, 4, and 3 respectively. Protein CMe has been found to be identical with a previously described trypsin inhibitor. The three proteins have been quantified in the different genetic stocks by HPLC. The different high-lysine mutations have different effects on the expression patterns of the three genes: CMe is markedly decreased and CMa and CMb are increased in mutant Risø 1508, whereas all three proteins are decreased in Risø 527 and increased in Risø 7 with respect to the wild-type Bomi; CMa and CMb are increased and CMe is unaffected in mutant Risø 56 with respect to the wild-type Carlsberg II; and protein CMe is markedly decreased in Hiproly barley as compared with its sister line CI4362. The implications of these results in connection with the evolution of CM proteins and with the characterization of high-lysine mutations are discussed.

Amino Acid Sequence↗
PubMed1985

The mode of action of restrictocin and mitogillin on eukaryotic ribosomes. Inhibition of brain protein synthesis, cleavage and sequence of the ribosomal RNA fragment.

The relationship between the process of rat brain protein synthesis inhibition by restrictocin and mitogillin and the induction of a specific cleavage in 28S rRNA has been examined. Restrictocin or mitogillin at a concentration of 6 nM inhibits protein synthesis to a level of 80%. The inhibition induced by 6 nM alpha-sarcin is of the same order, indicating that those molecules all catalytically inhibit protein synthesis. When restrictocin or mitogillin was reduced and alkylated, a 100-fold higher concentration of these chemically modified molecules was needed in order to obtain the same inhibition as with native molecules, suggesting a close correlation between activity and configuration of the inhibitors. The inhibitory activity of restrictocin or mitogillin was completely lost after oxidation with performic acid. The inhibition of protein synthesis was always correlated with the production of a fragment from the 3'-end of the 28S rRNA. This rRNA fragment had the same electrophoretic mobility as that produced by alpha-sarcin. The 5'-end sequence of the rRNA fragment produced by restrictocin, mitogillin or alpha-sarcin is AGGAA, demonstrating that restrictocin and mitogillin inhibit protein synthesis in the same way as does alpha-sarcin. The conservation of this region of the rRNA from archaebacteria to eukaryotic ribosomes indicates its importance in the elongation process.

Alkylation↗
PubMed1985

Structure and mode of action of microcin 7, an antibacterial peptide produced by Escherichia coli.

Microcin 7 is a small peptide produced and excreted to the culture medium by stationary-phase Escherichia coli cells harboring the pMccC7 plasmid (formerly named pRYC7). This peptide inhibited the growth of the enterobacteria phylogenetically closer to E. coli, apparently by blocking protein biosynthesis. The molecule was degraded with trypsin, and the resulting fragments were purified and sequenced. The results show that microcin 7 is a linear heptapeptide blocked at both ends.

Acylation↗
PubMed1983

Isolation of human complex-forming glycoprotein, heterogeneous in charge (protein HC), and its IgA complex from plasma. Physiochemical and immunochemical properties, normal plasma concentration.

Human complex-forming glycoprotein, heterogeneous in charge (protein HC) has previously been isolated from urine and immunochemically shown to be present in low and high molecular weight forms in blood plasma (Tejler, L., and Grubb, A. O. (1976) Biochim. Biophys. Acta 439, 82-94). In the present work, the major low and high molecular weight forms of the protein were isolated from plasma by immunosorption followed by gel chromatography. The plasma low molecular weight protein HC and the urinary protein had similar, if not identical, molecular weight, amino acid composition, NH2-terminal and carboxyl-terminal amino acid sequences and electrophoretic mobility. The low molecular weight plasma protein HC carried a yellow chromophore like the urinary protein, but its molar extinction coefficient at 280 nm was lower and its charge heterogeneity less pronounced than that of urinary protein HC. The plasma high molecular weight protein HC had a hydrodynamic volume which was greater than that of monomeric IgA but smaller than that of dimeric IgA. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the isolated high molecular weight protein followed by electrophoretic blotting and immunochemical analysis demonstrated that the protein contained four polypeptide chains: two light immunoglobulin chains (Mr = 23,000), one IgA alpha-chain (Mr = 54,000), and one chain with Mr approximately 90,000 which carried both alpha-chain and protein HC antigenic determinants. Whether the protein HC X IgA complex is a functionally significant part of the humoral immune system cannot be decided without further experimentation, but the complex was found to be completely absent from the blood plasma of patients with a selective deficiency of IgA-secreting immunocytes. The isolated low and high molecular weight plasma protein HC components were used as standard proteins in the construction of a quantitative crossed immunoelectrophoretic assay for the simultaneous quantitation of the two major protein HC components in blood plasma. The plasma concentrations of the low and high molecular weight protein HC components were measured by this method in 13 healthy Caucasians. The results for the low molecular weight protein HC were: mean, 20.3 mg/liter, S.D., 3.2 mg/liter, range, 13.6-26.0 mg/liter; and for the protein HC X IgA complex: mean, 293 mg/liter, S.D., 176 mg/liter, range, 36-620 mg/liter.

Adult↗
PubMed1983

The primary structure of human serum transferrin. The structures of seven cyanogen bromide fragments and the assembly of the complete structure.

The amino acid sequences of seven cyanogen bromide fragments of human serum transferrin have been determined, and the primary structure of transferrin established by determining the order of these and three additional fragments (Sutton, M. R., MacGillivray, R. T. A., and Brew, K. (1975) Eur. J. Biochem. 51, 43-48) in the polypeptide chain. The order of the fragments was deduced from peptides that overlap methionyl residues which were obtained by thermolysin digestion of performic acid-oxidized transferrin or by partial peptic hydrolysis of unmodified transferrin, together with other evidence. The polypeptide chain of transferrin contains 679 amino acid residues, which together with the two N-linked oligosaccharide chains gives a calculated molecular weight of 79,570. Transferrin consists of two homologous domains (residues 1-336, 337-679), each associated with a single Fe-binding site, with both sites of glycosylation in the carboxyl-terminal domain at positions 413 and 611. Consideration of the primary structure in relation to previously published results provides information concerning the evolutionary development of transferrins and related proteins, and the locations of metal-binding residues in the transferrin molecule.

Amino Acid Sequence↗