A Cutinase from Trichoderma reesei with a Lid-Covered Active Site and Kinetic Properties of True Lipases

被引:45
作者
Roussel, Alain [1 ,2 ]
Amara, Sawsan [3 ]
Nyyssola, Antti [4 ]
Mateos-Diaz, Eduardo [3 ]
Blangy, Stephanie [1 ,2 ]
Kontkanen, Hanna [4 ]
Westerholm-Pantinen, Ann [4 ]
Carriere, Frederic [3 ]
Cambillau, Christian [1 ,2 ]
机构
[1] Aix Marseille Univ, Architecture & Fonct Macromol Biol, F-13284 Marseille 09, France
[2] CNRS, UMR7257, Architecture & Fonct Macromol Biol, F-13288 Marseille 09, France
[3] Aix Marseille Univ, UMR7282, CNRS, F-13402 Marseille 20, France
[4] VTT Biotechnol, FIN-02044 Espoo, Finland
关键词
cutinase; lipase; Trichoderma reesei; X-ray structure; interfacial activation; FUSARIUM-SOLANI CUTINASE; CRYSTAL-STRUCTURE; PANCREATIC LIPASE; INTERFACIAL ACTIVATION; GASTRIC LIPASE; CELLULASE PRODUCTION; LIPOLYTIC ENZYME; TRIAD FORMS; X-RAY; COMPLEX;
D O I
10.1016/j.jmb.2014.09.003
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
Cutinases belong to the alpha/beta-hydrolase fold family of enzymes and degrade cutin and various esters, including triglycerides, phospholipids and galactolipids. Cutinases are able to degrade aggregated and soluble substrates because, in contrast with true lipases, they do not have a lid covering their catalytic machinery. We report here the structure of a cutinase from the fungus Trichoderma reesei (Tr) in native and inhibitor-bound conformations, along with its enzymatic characterization. A rare characteristic of Tr cutinase is its optimal activity at acidic pH. Furthermore, Trcutinase, in contrast with classical cutinases, possesses a lid covering its active site and requires the presence of detergents for activity. In addition to the presence of the lid, the core of the Tr enzyme is very similar to other cutinase cores, with a central five-stranded beta-sheet covered by helices on either side. The catalytic residues form a catalytic triad involving Ser164, His229 and Asp216 that is covered by the two N-terminal helices, which form the lid. This lid opens in the presence of surfactants, such as p-octylglucoside, and uncovers the catalytic crevice, allowing a C11Y4 phosphonate inhibitor to bind to the catalytic serine. Taken together, these results reveal Tr cutinase to be a member of a new group of lipolytic enzymes resembling cutinases but with kinetic and structural features of true lipases and a heightened specificity for long-chain triglycerides. (C) 2014 Elsevier Ltd. All rights reserved.
引用
收藏
页码:3757 / 3772
页数:16
相关论文
共 54 条
[1]   CRYSTALLIZATION AND PRELIMINARY-X-RAY STUDY OF A RECOMBINANT CUTINASE FROM FUSARIUM-SOLANI-PISI [J].
ABERGEL, C ;
MARTINEZ, C ;
FONTECILLACAMPS, J ;
CAMBILLAU, C ;
DEGEUS, P ;
LAUWEREYS, M .
JOURNAL OF MOLECULAR BIOLOGY, 1990, 215 (02) :215-216
[2]   The galactolipase activity of some microbial lipases and pancreatic enzymes [J].
Amara, Sawsan ;
Lafont, Dominique ;
Parsiegla, Goetz ;
Point, Vanessa ;
Chabannes, Aurelie ;
Rousset, Audric ;
Carriere, Frederic .
EUROPEAN JOURNAL OF LIPID SCIENCE AND TECHNOLOGY, 2013, 115 (04) :442-451
[3]   Continuous measurement of galactolipid hydrolysis by pancreatic lipolytic enzymes using the pH-stat technique and a medium chain monogalactosyl diglyceride as substrate [J].
Amara, Sawsan ;
Lafont, Dominique ;
Fiorentino, Brice ;
Boullanger, Paul ;
Carriere, Frederic ;
De Caro, Alain .
BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR AND CELL BIOLOGY OF LIPIDS, 2009, 1791 (10) :983-990
[4]   THE CCP4 SUITE - PROGRAMS FOR PROTEIN CRYSTALLOGRAPHY [J].
BAILEY, S .
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY, 1994, 50 :760-763
[5]   Refinement of severely incomplete structures with maximum likelihood in BUSTER-TNT [J].
Blanc, E ;
Roversi, P ;
Vonrhein, C ;
Flensburg, C ;
Lea, SM ;
Bricogne, G .
ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY, 2004, 60 :2210-2221
[6]   A SERINE PROTEASE TRIAD FORMS THE CATALYTIC CENTER OF A TRIACYLGLYCEROL LIPASE [J].
BRADY, L ;
BRZOZOWSKI, AM ;
DEREWENDA, ZS ;
DODSON, E ;
DODSON, G ;
TOLLEY, S ;
TURKENBURG, JP ;
CHRISTIANSEN, L ;
HUGEJENSEN, B ;
NORSKOV, L ;
THIM, L ;
MENGE, U .
NATURE, 1990, 343 (6260) :767-770
[7]   A MODEL FOR INTERFACIAL ACTIVATION IN LIPASES FROM THE STRUCTURE OF A FUNGAL LIPASE-INHIBITOR COMPLEX [J].
BRZOZOWSKI, AM ;
DEREWENDA, U ;
DEREWENDA, ZS ;
DODSON, GG ;
LAWSON, DM ;
TURKENBURG, JP ;
BJORKLING, F ;
HUGEJENSEN, B ;
PATKAR, SA ;
THIM, L .
NATURE, 1991, 351 (6326) :491-494
[8]   Pancreatic lipase structure-function relationships by domain exchange [J].
Carriere, F ;
Thirstrup, K ;
Hjorth, S ;
Ferrato, F ;
Nielsen, PF ;
WithersMartinez, C ;
Cambillau, C ;
Boel, E ;
Thim, L ;
Verger, R .
BIOCHEMISTRY, 1997, 36 (01) :239-248
[9]   Inhibition of human gastric and pancreatic lipases by chiral alkylphosphonates.: A kinetic study with 1,2-didecanoyl-sn-glycerol monolayer [J].
Cavalier, JF ;
Ransac, S ;
Verger, R ;
Buono, G .
CHEMISTRY AND PHYSICS OF LIPIDS, 1999, 100 (1-2) :3-31
[10]   How gastric lipase, an interfacial enzyme with a Ser-His-Asp catalytic triad, acts optimally at acidic pH [J].
Chahinian, H ;
Snabe, T ;
Attias, C ;
Fojan, P ;
Petersen, SB ;
Carrière, F .
BIOCHEMISTRY, 2006, 45 (03) :993-1001