Thermostable aminopeptidase from Pyrococcus horikoshii

被引:36
作者
Ando, S
Ishikawa, K
Ishida, H
Kawarabayasi, Y
Kikuchi, H
Kosugi, Y
机构
[1] Natl Inst Biosci & Human Technol, Tsukuba, Ibaraki 3058566, Japan
[2] Natl Inst Technol & Evaluat, Tokyo 1510066, Japan
关键词
aminopeptidase; thermostability; Archaeon; Pyrococcus horikoshii;
D O I
10.1016/S0014-5793(99)00257-4
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
From the genome sequence data of the thermophilic archaeon Pyrococcus horikoshii, an open reading frame was found which encodes a protein (332 amino acids) homologous with an endoglucanase from Clostridium thermocellum (42% identity), deblocking aminopeptidase from Pyrococcus furiosus (42% identity) and an aminopeptidase from Aeromonas proteolytica (18% identity). This gene was cloned and expressed in Escherichia coli, and the characteristics of the expressed protein were examined. Although endoglucanase activity was not detected, this protein was found to have aminopeptidase activity to cleave the N-terminal amino acid from a variety of substrates including both N-blocked and non-blocked peptides, The enzyme was stable at 90 degrees C, with the optimum temperature over 90 degrees C. The metal ion bound to this enzyme was calcium, but it was not essential for the aminopeptidase activity. Instead, this enzyme required the cobalt ion for activity. This enzyme is expected to be useful for the removal of N(alpha)-acylated residues in short peptide sequence analysis at high temperatures. (C) 1999 Federation of European Biochemical Societies.
引用
收藏
页码:25 / 28
页数:4
相关论文
共 13 条
[1]   BASIC LOCAL ALIGNMENT SEARCH TOOL [J].
ALTSCHUL, SF ;
GISH, W ;
MILLER, W ;
MYERS, EW ;
LIPMAN, DJ .
JOURNAL OF MOLECULAR BIOLOGY, 1990, 215 (03) :403-410
[2]   CRYSTAL-STRUCTURE OF AEROMONAS-PROTEOLYTICA AMINOPEPTIDASE - A PROTOTYPICAL MEMBER OF THE CO-CATALYTIC ZINC ENZYME FAMILY [J].
CHEVRIER, B ;
SCHALK, C ;
DORCHYMONT, H ;
RONDEAU, JM ;
TARNUS, C ;
MORAS, D .
STRUCTURE, 1994, 2 (04) :283-291
[3]  
DOI E, 1981, ANAL BIOCHEM, V118, P173, DOI 10.1016/0003-2697(81)90175-5
[4]  
GADE W, 1978, J BIOL CHEM, V253, P5012
[5]   Pyrococcus horikoshii sp. nov., a hyperthermophilic archaeon isolated from a hydrothermal vent at the Okinawa Trough [J].
Gonzalez, JM ;
Masuchi, Y ;
Robb, FT ;
Ammerman, JW ;
Maeder, DL ;
Yanagibayashi, M ;
Tamaoka, J ;
Kato, C .
EXTREMOPHILES, 1998, 2 (02) :123-130
[6]   KINETICS OF HYDROLYTIC REACTION CATALYZED BY CRYSTALLINE BACTERIAL ALPHA-AMYLASE .3. THE INFLUENCE OF TEMPERATURE [J].
HIROMI, K ;
TAKASAKI, Y ;
ONO, S .
BULLETIN OF THE CHEMICAL SOCIETY OF JAPAN, 1963, 36 (05) :563-569
[7]   SITE-DIRECTED MUTAGENESIS BY OVERLAP EXTENSION USING THE POLYMERASE CHAIN-REACTION [J].
HO, SN ;
HUNT, HD ;
HORTON, RM ;
PULLEN, JK ;
PEASE, LR .
GENE, 1989, 77 (01) :51-59
[8]   Acylamino acid-releasing enzyme from the thermophilic archaeon Pyrococcus horikoshii [J].
Ishikawa, K ;
Ishida, H ;
Koyama, Y ;
Kawarabayasi, Y ;
Kawahara, J ;
Matsui, E ;
Matsui, I .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1998, 273 (28) :17726-17731
[9]  
Kamp RM, 1998, J PROTEIN CHEM, V17, P512
[10]  
Kawarabayasi Y, 1998, DNA Res, V5, P147, DOI 10.1093/dnares/5.2.147