Finding a putative enzyme belonging to amidohydrolase family from Pyrococcus furiosus based on the conserved motifs = 보존부위에 근거한 Pyrococcus furiosus로부터의 새로운 amidohydrolase family 효소의 탐색 DP BS

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Nowadays, for identification of the function and structure prediction of theprotein being product of new gene, comparison of sequence homology by computational analysis between this new and already known protein seems to be useful tool. These works are mainly based on the conservation of structure concerning the divergent sequence and on the insight of the similar catalytic activity over various substrates. Based on the analysis of conservation pattern in the structure and similarity in the catalytic activity of the enzymes, we have proposed a cyclic amidohydrolase family, including D-hydantoinase, dihydropyrimidinase, dihydro-orotase, allantoinase. We found a number of highly conserved regions and amino acid residues. It is suggested that in archaea, there were existences of thermostable counterparts to known classes enzymes previously characterized from mesophilic sources, which may be concerned to same crucial enzyme reaction in vivo. There was some trial to detect an additional member of amidohydrolase family by computational analysis. Therefore, we attempt to find the new enzyme belonging to this family from the partial available genome databases of Pyrococcus furiosus by using of the homology epitope deduced from amidohydrolase family. Based on these four regions, we probed the whole genome sequence of P.furiosus and led to isolation of 1.5kb ORF fragment encoding putative enzyme, PNT. The ORF encoding putative enzyme was obtained from genomic DNA of P.furiosus, and this 1.5kb fragment was cloned into the plasmid vector pTrc99A, pMAL via EcoRI, PstI restriction sites. and expressed in E Coli. The expressed protein was isolated and its enzymatic activity was tested. Based on the similarities in the conserved region between putative enzyme and member of the amidohydrolase family, we suggested an evolutionary relationship and functional identification
Kim, Hak-Sungresearcher김학성researcher
한국과학기술원 : 생물과학과,
Issue Date
158717/325007 / 000983391

학위논문(석사) - 한국과학기술원 : 생물과학과, 2000.2, [ iv, 43 p. ]


Pyrococcus furiosus; Conserved regions; Amidohydrolase family; Putative enzyme; 파이로코커스 퓨리오서스; 아미도하이드롤에이즈 패밀리; 새로운 효소; 보존 부위

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