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Biomedical subjects

D Mu

Publications and source records attributed to D Mu.

30 records · Page 2Linked to original sources
PubMed1996

[The treated effects of nimodipine on calcium of erythrocyte in experimental hypoxic-ischemic encephalopathy].

An animal model of hypoxic-ischemic encephalopathy (HIE) in newborn pigs was set up to study the histopathological changes of brain injury by light- and electron-microscopy, the changes in total calcium content and calcium iron of RBC, and the effects of two different doses of Nimodipine on HIE. Sixty-one newborn pigs were randomly divided into seven groups, the normal control group (n = 9); the HIE 1-hour group (n = 8) and 24-hour group (n = 8); HIE+ Nimodipine (1 microgram/kg.min-1) 1-hour group (n = 8) and 24-hour group (n = 8). The results showed that these were apparent brain edema, hematencephalon, neuron necrosis, and intracellular body degeneration in the HIE 1-hour group. The levels of RBC TCa and RBC Cai2+ in the HIE 1-hour group were significantly higher than those in the normal control group (P < 0.01). The brain tissues in all groups treated with Nimodipine showed histopathological betterment Compared with the HIE groups. The levels of RBC TCa and RBC Cai2+ in the Nimodipine treatment groups were significantly lower than those in HIE groups (P < 0.01) and were close to the levels in the normal control group (P < 0.05). The authors suggested that early application of Nimodipine in case of postnatal asphyxia might protect the newborn from severe hypoxicischemic brain lesions.

Animals↗
PubMed1995

Human DNA repair excision nuclease. Analysis of the roles of the subunits involved in dual incisions by using anti-XPG and anti-ERCC1 antibodies.

Human DNA repair excision nuclease removes DNA damage by incising on both sides of the lesion in a precise manner. The activity requires participation of 16-17 polypeptides. Of these, the XPF.ERCC1 complex and XPG were predicted to carry the nuclease active sites based on studies with the recombinant proteins and the yeast homologs of these proteins. Furthermore, recent work with model (undamaged) substrates have led to predictions of the roles of these proteins in incising 5' or 3' to the lesion. We have used damaged DNA substrates and antibodies to XPG and ERCC1 to test these predictions. Our results reveal that anti-XPG antibodies change the site of 3' incision and at high concentration inhibit the 3' incision without significantly affecting the 5' incision, indicating that XPG makes the 3' incision and further that under this condition 5' incision can occur without 3' incision. In contrast, anti-ERCC1 antibodies inhibit both the 3' and 5' incisions. Using a defined system for excision repair we also demonstrate that the 3' incision can occur without the 5' incision, leading us to conclude that under certain conditions the two incisions can occur independently.

Animals↗
PubMed1995

The general transcription-repair factor TFIIH is recruited to the excision repair complex by the XPA protein independent of the TFIIE transcription factor.

Recent studies have revealed that the general transcription factor TFIIH is also a general excision repair factor which, along with several other proteins, is required for transcription-independent excision reaction. As a general transcription factor, TFIIH is recruited to RNA polymerase II-promoter complex by another general transcription factor called TFIIE. We were interested in knowing whether TFIIE is also involved in recruiting TFIIH to the excision repair complex. We found that cell-free extract depleted of TFIIE carried out excision repair at a normal rate, leading us to conclude that TFIIE is not involved in recruiting TFIIH to the damage site and has no role in general excision repair. In contrast, the human damage recognition protein XPA specifically binds to TFIIH and apparently recruits it to the damage site. The carboxyl-terminal half of XPA is responsible for specific interaction with TFIIH. The C261S/C264S mutant of XPA bound the ERCC1-XPF complex normally, but failed to bind TFIIH and failed to complement an XP-A mutant cell-free extract indicating that the XPA-TFIIH interaction is essential to effecting the excision reaction. Interestingly, XPA also binds to the p34 subunit of TFIIE specifically and in competition with the p56 subunit of TFIIE. This latter interaction has no apparent role in general excision repair but may be relevant in the transcription-coupled repair reaction.

Amino Acid Sequence↗
PubMed1995

Reconstitution of human DNA repair excision nuclease in a highly defined system.

Xeroderma pigmentosum is a hereditary disease caused by defective DNA repair. Somatic cell genetics and biochemical studies with cell-free extracts indicate that at least 16 polypeptides are required to carry out the repair reaction proper, i.e. the removal of the lesion from the DNA by the dual incisions of the damaged strand. To find out if these proteins are necessary and sufficient for excision repair, they were obtained at a high level of purity in five fractions. The mixture of these five fractions reconstituted the excision nuclease (excinuclease) activity. Using the reconstituted excinuclease, we found that the excised fragment remains associated with the post-incision DNA-protein complex, suggesting that accessory proteins are needed to release the excised oligomer.

Animals↗
PubMed1994

Human and E.coli excinucleases are affected differently by the sequence context of acetylaminofluorene-guanine adduct.

Synthetic DNA substrates containing an acetylaminofluorene (AAF) adduct at each of the three guanine in the G1G2CG3CC sequence were constructed and tested as substrates for reconstituted E.coli (A)BC excinuclease and human excinuclease in HeLa cell-free extract (CFE). The (A)BC excinulcease repaired the three substrates with relative efficiencies of G1:G2:G3 of 100:18:66 in agreement with an earlier report [Seeberg, E., and Fuchs, R.P.P. (1990) Proc. Natl Acad. Sci. USA 87, 191-194]. The same lesions were repaired by the human excinuclease with the strikingly different efficiencies of G1:G2:G3 as 38:100:68. These results reveal that the human excinuclease is affected by the sequence context of the lesion in a different manner than its prokaryotic counterpart.

Base Sequence↗
PubMed1994

Primary structures for a mammalian cellular and serum copper amine oxidase.

The 6-hydroxydopa quinone-containing active site peptide from bovine serum amine oxidase has been found to be highly homologous to a segment of a cloned human kidney amiloride-binding protein (Barbry, P., Champe, M., Chassande, O., Munemitsu, S., Champigny, G., Lingueglia, E., Maes, P., Frelin, C., Tartar, A., Ullrich, A., and Lazdunski, M. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 7347-7351). Additionally, a second 38-residue tryptic peptide (peptide XI) isolated from bovine serum amine oxidase shows 82% identity with a portion near the carboxyl terminus of the human kidney amiloride-binding protein. When an extended active site peptide was isolated from porcine kidney diamine oxidase (Janes, S. M., Palcic, M. M., Scaman, C. H., Smith, A. J., Brown, D. E., Dooley, D. M., Mure, M., and Klinman, J. P. (1992) Biochemistry 31, 12147-12154), it was found to be fully contained in the human kidney amiloride-binding protein. Examination of amiloride binding to bovine serum amine oxidase and porcine kidney diamine oxidase reveals dissociation constants of 196 and 9.1 microM, respectively. Taken together, these findings indicate that the cDNA isolated for human kidney amiloride-binding protein encodes a human kidney diamine oxidase. Two oligonucleotides, based on the tryptic peptide XI and active-site peptide of bovine serum amine oxidase, were used to amplify a portion of cDNA from a commercial bovine liver cDNA library through the use of the polymerase chain reaction. A full-length clone (2.7 kilobase pairs) for bovine serum amine oxidase was subsequently obtained through screening of the same cDNA library with the amplified 0.7-kilobase pair cDNA. These studies provide the first primary sequences for a mammalian cellular and serum copper amine oxidase. Computer alignment of amine oxidase cDNA-derived protein sequences reveals three conserved histidine residues, which are likely to be ligands to copper.

Amine Oxidase (Copper-Containing)↗
PubMed1992

Tyrosine codon corresponds to topa quinone at the active site of copper amine oxidases.

The recently discovered organic cofactor of bovine serum amine oxidase, topa quinone, is an uncommon amino acid residue in the polypeptide backbone (Janes, S. M., Mu, D., Wemmer, D., Smith, A. J., Kaur, S., Maltby, D., Burlingame, A. L., and Klinman, J. P. (1990) Science 248, 981-987). The amine oxidase gene from the yeast Hansenula polymorpha has been cloned and sequenced (Bruinenberg, P. G., Evers, M., Waterham, H. R., Kuipers, J., Arnberg, A. C., and Geert, A. B. (1989) Biochim. Biophys. Acta 1008, 157-167). In order to understand the incorporation of topa quinone in eukaryotes, we have isolated yeast amine oxidase from H. polymorpha. Following protocols established with bovine serum amine oxidase, yeast amine oxidase was derivatized with [14C]phenylhydrazine, followed by thermolytic digestion and isolation of a dominant radiolabeled peptide by high pressure liquid chromatography. Comparison of resonance Raman spectra for this peptide to spectra of a model compound demonstrates that topa quinone is the cofactor. By alignment of a DNA-derived yeast amine oxidase sequence with the topa quinone-containing peptide sequence, it is found that the tyrosine codon, UAC, corresponds to topa quinone in the mature protein. In a similar manner, alignment of a tryptic peptide from bovine serum amine oxidase implicates tyrosine as the precursor to topa quinone in mammals.

Amine Oxidase (Copper-Containing)↗
PubMed1991

The organic functional group in copper-containing amine oxidases. Resonance Raman spectra are consistent with the presence of topa quinone (6-hydroxydopa quinone) in the active site.

Resonance Raman spectroscopy has been used to probe the structure of the organic cofactor in copper-containing amine oxidases from bovine plasma, porcine kidney, pea seedlings, and the bacterium Arthrobacter P1. The enzymes were first derivatized with phenylhydrazine or p-nitrophenylhydrazine; resonance Raman spectra were obtained on the intact derivatized enzymes and on a derivatized active-site peptide isolated from bovine plasma amine oxidase. Spectra of the intact amine oxidase phenylhydrazones are practically identical, consistent with the enzymes examined containing a similar cofactor. Only minor frequency shifts and some intensity variations are detected between the resonance Raman spectra of intact bovine plasma amine oxidase and the isolated peptide. These spectral perturbations are attributable to differences in the micro-environment between the intact, folded protein and the isolated small peptide in aqueous solution. This rules out the possibility that a new structure is formed during the isolation of the derivatized active-site peptide. Importantly, the resonance Raman spectra of the phenylhydrazine and p-nitrophenylhydrazine derivatives of the bovine plasma amine oxidase peptide are identical to the spectra of the corresponding derivatives of topa quinone (6-hydroxydopa quinone). Hence these data provide strong, independent support for the recent identification of topa as the organic functional group in bovine plasma amine oxidase (Janes, S. M., Mu, D., Wemmer, D., Smith, A. J., Kaur, S., Maltby, D., Burlingame, A. L., and Klinman, J.P. (1990) Science 248, 981-987).

Amine Oxidase (Copper-Containing)↗
PubMed1990

A new redox cofactor in eukaryotic enzymes: 6-hydroxydopa at the active site of bovine serum amine oxidase.

An active site, cofactor-containing peptide has been obtained in high yield from bovine serum amine oxidase. Sequencing of this pentapeptide indicates: Leu-Asn-X-Asp-Tyr. Analysis of the peptide by mass spectrometry, ultraviolet-visible spectroscopy, and proton nuclear magnetic resonance leads to the identification of X as 6-hydroxydopa. This result indicates that, contrary to previous proposals, pyrroloquinoline quinone is not the active site cofactor in mammalian copper amine oxidases. Although 6-hydroxydopa has been implicated in neurotoxicity, the data presented suggest that this compound has a functional role at an enzyme active site.

Amine Oxidase (Copper-Containing)↗