Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Evidence is presented for the uptake of the human X chromosome by human-Chinese hamster cell hybrids which lack H P R T activity, following incubation with isolated human HeLa S3 chromosomes. Sixteen independent clonal cell lines were isolated in H A T medium, all of which contained a human X chromosome as determined by trypsin-Giemsa staining. The frequency of H A T-resistant clones was 32 x 10(-6) when 10(7) cells were incubated with 10(8) HeLa chromosomes. Potential reversion of the hybrid cells in H A T medium was less than 5 x 10(-7). The 16 isolated cell lines all contained activity of the human X-linked marker enzymes H P R T, P G K,alpha-Gal A, and G6PD, as determined by electrophoresis. The phenotype of G6PD was G6PD A, corresponding to G6PD A in HeLa cells. The human parental cells used in the fusion to form the hybrids had the G6PD B phenotype. The recipient cells gave no evidence of containing human X chromosomes. These results indicate that incorporation and expression of HeLa X chromosomes is accomplished in human-Chinese hamster hybrids which lack a human X chromosome.
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PMID:Transfer of the human X chromosome to human--Chinese hamster cell hybrids via isolated HeLa metaphase chromosomes. 102 48

Testosterone-treated calf thymocytes produce increased amounts of proteins, termed lipokinins, that stimulate phospholipase A2 from snake venom and mammalian tissue. The induction of these proteins by testosterone is blocked by cycloheximide and, thus, requires new protein synthesis. These proteins activate phospholipase A2 stoichiometrically. They are inactivated by boiling, trypsin or alkaline phosphatase but not by deoxyribonuclease or ribonuclease. Lipokinins significantly repair the failure of masculinization in the Tfm mouse with an X-linked deficiency of androgen-receptor. Thus, the post-receptor effects of testosterone on embryonic genitalia may be mediated through stimulation of phospholipase A2 by lipokinins. Moreover, lipokinins may be involved as stimulators of the arachidonic acid cascade, as lipocortins are inhibitors.
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PMID:John Lattimer lecture. Lipokinins: novel phospholipase A2 activators mediate testosterone effects on embryonic genitalia. 318 94

The In(Lu) gene, which inhibits the expression of Lutheran blood group antigens by red cells (RBCs), also down-regulates the expression of an 80-kD glycoprotein, In(Lu)-related p80, by both RBCs and a subset of white cells. This study examined the expression of multiple-RBC p80 epitopes by autosomal and X-linked recessive-type Lu(a-b-) RBCs in order to explore the relationship, if any, between expression of In(Lu)-related p80 and Lutheran antigens. Both autosomal and X-linked types of recessive Lu(a-b-) RBCs expressed near-normal to increased amounts of p80 antigens, as measured by radioimmunoassay. P80 from both types of recessive Lu(a-b-) RBCs had apparently normal molecular weight in denaturing polyacrylamide gels and showed normal sensitivity to digestion by trypsin and chymotrypsin. Thus, the absence of Lutheran antigens on recessive-type Lu(a-b-) RBCs is not associated with decreased or absent p80 antigens. Furthermore, the XS2 gene probably does not act via a mechanism similar to that of the In(Lu) gene, since the expression of p80 remains undiminished in X-linked recessive-type Lu(a-b-) RBCs.
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PMID:Human red cell antigens. V. Expression of In(Lu)-related p80 antigens by recessive-type Lu(a-b-) red cells. 342 Jun 70

For comparative studies we have used the somatic cell hybridization approach to regionally map genes on the mouse X chromosome. Fibroblasts from a mouse with the balanced reciprocal translocation T(XD;16B5)16H were fused with a Chinese hamster cell line (V79/380-6) deficient in activity of the enzyme hypoxanthine phosphoribosyltransferase (HPRT). Interpecific cell hybrids were initially selected for retention of the mouse translocation chromosome carrying the Hprt gene. Subsequently, hybrid clones were counterselected to force segregation of this chromosome. Selected and counterselected hybrid clones were analyzed for their chromosome content by trypsin/Giemsa banding and for expression of the mouse forms of the X-linked enzymes HPRT and alpha-galactosidase (GALA) by isoelectric focusing. The results indicate that the breakpoint on the mouse X chromosome (in band XD) has separated the genes for HPRT (Hprt) and for GALA (Ags). Hprt is proximal to the breakpoint in region Xcen-XD and Ags is distal in region XD-Xter. The gene order in the mouse (centromere-Hprt-Ags) is therefore inverted when compared to the order of the homologous loci on the long arm of the human X (centromere-GALA-HPRT).
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PMID:Comparative gene mapping: order of loci on the X chromosome is different in mice and humans. 625 72

Monoclonal antibodies were raised against surface determinants of Streptococcus pneumoniae by hyperimmunizing X-linked immunodeficient (xid) CBA/N mice with the heat-killed rough strain R36A. 17 hybridomas produced antibody that bound intact R36A and did not cross-react with phosphocholine, an antigen common in the cell wall of all S. pneumoniae. The antibody produced by at least two of these hybridomas, Xi64 (IgM) and Xi126 (IgG2b), could protect mice from a lethal intravenous challenge of type 3 S. pneumoniae strains WU2 and A66 and of the type 2 strain D39. The minimum amount of antibody required to protect xid mice from 100 WU2 was 4.5 micrograms/mouse for Xi64 and 2.6 micrograms/mouse for Xi126,. Free phosphocholine, C-polysaccharide, and type 3 capsular polysaccharide all failed to inhibit the binding of Xi64 or Xi126 to R36A. These antibodies appeared to bind surface polypeptides, since treatment of R36A with either pepsin or trypsin, or of R36A lysate with trypsin, effectively eliminated the ability of Xi64 and Xi126 to bind antigens in these preparations. Binding studies indicated that these two antibodies recognized different epitopes that were expressed on several but not all serotypes of pneumococci.
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PMID:Monoclonal antibodies against protease-sensitive pneumococcal antigens can protect mice from fatal infection with Streptococcus pneumoniae. 638 34

The binding of the calcium antagonist [3H] nitrendipine ([3H] NDP) to brain and heart is described and the brain site is characterized. The binding is saturable, specific and of very high affinity with KD values of 0.16 nM in brain and 0.21 nM in heart. Our kinetic results are similar to those recently reported by two other groups (1, 2), indicating a saturable, high affinity binding site in brain. In brain the binding sites are enriched in crude nuclear and synaptosomal fractions. The highest levels of binding are seen in the hippocampus, caudate and cerebral cortex with much lower levels in the cerebellum and pons. Calcium has a marked stimulatory effect on [3H] NDP binding at 10(-4) M. Addition of 0.5 mM CaCl2 to EDTA treated membranes nearly doubles the number of binding sites. Of the many drugs and neurotransmitters tested only other calcium antagonists, i.e. verapamil, inhibit binding (IC50 = 250 nM). The inhibition of [3H] NDP binding by verapamil is apparently non-competitive and not complete, suggesting that [3H] NDP binds to several sites, only some of which are inhibited by verapamil. The [3H] NDP binding site is probably a protein since it is very sensitive to trypsin, heat and sulfhydryl reagents.
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PMID:Specific calcium antagonist binding sites in brain. 714 41

The current study aims to ascertain the fate of the melanocyte stimulating hormone (MSH) receptor and its ligand [Nle4, D-Phe7] alpha-MSH (NDP-MSH) following binding to murine B16 melanoma cells. Cells were incubated with [125I]-NDP-MSH for up to 180 min and binding, internalization and degradation determined. Intracellular trafficking of the radiolabel was assessed using Percoll density gradient centrifugation of homogenized cells. Receptor down-regulation and receptor mRNA levels were also measured over 96 hr after exposure to 1 microM ligand. NDP-MSH accumulation increased with time in a temperature-dependent manner and was inhibited by excess peptide. The ligand was rapidly internalized and translocated to the lysosomal compartment where it was degraded. Internalization was accompanied by a loss or down-regulation of cell surface receptors, suggesting internalization of the NDP-MSH-receptor complex. No recycling of the receptors between the plasma membrane and intracellular compartments could be detected in this cell-line. Approximately 15% of the surface receptors were resistant to down-regulation, possibly indicating receptor heterogeneity. Down-regulation persisted for up to 96 hr and was accompanied by a decrease in MSH receptor mRNA levels 48 hr after treatment. However, before this time, transcript levels were the same in treated and control cells. In contrast to what was seen with NDP-MSH, cell surface receptors removed with trypsin were rapidly replaced. These results show that NDP-MSH not only induced MSH receptor internalization but also inhibited receptor turnover, resulting in a prolonged down-regulation. It is concluded that, in B16 cells, the MSH receptor undergoes ligand-dependent internalization, resulting in a prolonged down-regulation.
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PMID:Binding and internalization of the melanocyte stimulating hormone receptor ligand [Nle4, D-Phe7] alpha-MSH in B16 melanoma cells. 902 81

Dosage compensation for X-chromosome-linked genes between male and female mammals occurs by inactivation of one of the two X chromosomes in the female. In somatic cells, either the paternal or the maternal X chromosome is randomly inactivated in a given cell. In contrast, in the extra-embryonic tissues of mice, the paternally-derived X chromosome is preferentially inactivated. The evidence for paternal X-chromosome inactivation in humans is controversial and remains to be clarified. In this study, we have developed a sensitive polymerase chain reaction (PCR) technique to investigate the methylation pattern of the X-linked androgen receptor (AR) gene. The 5' CpG island of this gene is methylated on the inactive X chromosome and hypomethylated on the active X chromosome in somatic cells. The paternal and the maternal alleles of the AR gene may be distinguished by a polymorphism in the number of CAG triplet repeats within the CpG island. As a source of human extra-embryonic tissue, we used chorionic villus (CV) samples from female conceptuses of 10-12 weeks gestation. From a tiny branch of a CV sample, two distinct cell lineages, the trophoblastic and mesodermal lineages, were dissected apart by trypsin digestion and micromanipulation and DNA was extracted separately from these purified tissues. Digestion of the DNA with the methylation-sensitive restriction enzyme, Hpall, followed by PCR amplification revealed that the paternal allele is preferentially methylated in trophoblastic cells, but not in mesodermal cells. These results strongly suggest that the paternal X chromosome is preferentially inactivated in the human extra-embryonic tissues early in development.
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PMID:Paternal X-chromosome inactivation in human trophoblastic cells. 923 11

The cDNA encoding bovine retinal isoforms of nucleoside diphosphate kinase (NDP-kinase, EC 2.7.4.6) has been cloned and sequenced. Based on the partial amino acid sequence of the enzyme determined after trypsin digestion of purified NDP-kinase, primers were synthesized and used to isolate two different cDNA clones encoding the full length of two NDP-kinase isoforms. The nucleotide sequences of these clones contained open reading frames encoding 152-residue polypeptides with calculated molecular masses of 17.262 and 17.299 kDa, similar to that determined for the subunits of purified enzyme (17.5 and 18.5 kDa). The deduced NDP-kinase sequences showed high similarity with the known NDP-kinase sequences from other sources.
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PMID:Isolation of cDNA clones encoding two isoforms of nucleoside diphosphate kinase from bovine retina. 1022 34

The PHEX gene that is mutated in patients with X-linked hypophosphatemia (XLH) encodes a protein homologous to the M13 family of zinc metallopeptidases. The present study was undertaken to assess the impact of nine PHEX missense mutations on cellular trafficking, endopeptidase activity, and protein conformation. Secreted forms of wild-type and mutant PHEX proteins were generated by PCR mutagenesis; these included C85R, D237G, Y317F, G579R, G579V, S711R, A720T, and F731Y identified in XLH patients, and E581V, which in neutral endopeptidase 24.11 abolishes catalytic activity but not plasma membrane localization. The wild-type and D237G, Y317F, E581V, and F731Y proteins were terminally glycosylated and secreted into the medium, whereas the C85R, G579R, G579V, S711R, and A720T proteins were trapped inside the transfected cells. Growing the cells at 26 C permitted the secretion of G579V, S711R, and A720T proteins, although the yield of rescued G579V was insufficient for further analysis. Endopeptidase activity of secreted and rescued PHEX proteins, assessed using a novel internally quenched fluorogenic peptide substrate, revealed that E581V and S711R are completely inactive; D237G and Y317F exhibit 50-60% of wild-type activity; and A720T and F731Y retain full catalytic activity. Conformational analysis by limited proteolysis demonstrated that F731Y is more sensitive to trypsin and D237G is more resistant to endoproteinase Glu-c than the wild-type protein. Thus, defects in protein trafficking, endopeptidase activity, and protein conformation account for loss of PHEX function in XLH patients harboring these missense mutations.
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PMID:Structure and function of disease-causing missense mutations in the PHEX gene. 1272 77


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