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Title: Structural insight into mechanism and diverse substrate selection strategy of L-ribulokinase

Abstract

The araBAD operon encodes three different enzymes required for catabolism of L-arabinose, which is one of the most abundant monosaccharides in nature. L-ribulokinase, encoded by the araB gene, catalyzes conversion of L-ribulose to L-ribulose-5-phosphate, the second step in the catabolic pathway. Unlike other kinases, ribulokinase exhibits diversity in substrate selectivity and catalyzes phosphorylation of all four 2-ketopentose sugars with comparable k{sub cat} values. To understand ribulokinase recognition and phosphorylation of a diverse set of substrates, we have determined the X-ray structure of ribulokinase from Bacillus halodurans bound to L-ribulose and investigated its substrate and ATP co-factor binding properties. The polypeptide chain is folded into two domains, one small and the other large, with a deep cleft in between. By analogy with related sugar kinases, we identified {sup 447}{und GG}LPQ{und K}{sup 452} as the ATP-binding motif within the smaller domain. L-ribulose binds in the cleft between the two domains via hydrogen bonds with the side chains of highly conserved Trp126, Lys208, Asp274, and Glu329 and the main chain nitrogen of Ala96. The interaction of L-ribulokinase with L-ribulose reveals versatile structural features that help explain recognition of various 2-ketopentose substrates and competitive inhibition by L-erythrulose. Comparison of our structure to that ofmore » the structures of other sugar kinases revealed conformational variations that suggest domain-domain closure movements are responsible for establishing the observed active site environment.« less

Authors:
; ;
Publication Date:
Research Org.:
BROOKHAVEN NATIONAL LABORATORY (BNL)
Sponsoring Org.:
ELI LILLY & COMPANY
OSTI Identifier:
1044005
Report Number(s):
BNL-96553-2012-JA
Journal ID: ISSN 0887-3585; PSFGEY; 600301010; TRN: US201214%%260
DOE Contract Number:  
DE-AC02-98CH10886
Resource Type:
Journal Article
Journal Name:
Proteins
Additional Journal Information:
Journal Volume: 80; Journal Issue: 1; Journal ID: ISSN 0887-3585
Country of Publication:
United States
Language:
English
Subject:
08 HYDROGEN; 59 BASIC BIOLOGICAL SCIENCES; ARABINOSE; BACILLUS; CATABOLISM; CHAINS; CLOSURES; CRYSTAL STRUCTURE; ENZYMES; HYDROGEN; MONOSACCHARIDES; NITROGEN; PHOSPHORYLATION; PHOSPHOTRANSFERASES; POLYPEPTIDES; RIBULOSE; SACCHARIDES; SACCHAROSE; SUBSTRATES; crystal structure; ribulokinase; ribulose; araBAD; araB; arabinose; catabolism

Citation Formats

Agarwal R., Swaminathan S., and Burley, S. K. Structural insight into mechanism and diverse substrate selection strategy of L-ribulokinase. United States: N. p., 2012. Web. doi:10.1002/prot.23202.
Agarwal R., Swaminathan S., & Burley, S. K. Structural insight into mechanism and diverse substrate selection strategy of L-ribulokinase. United States. doi:10.1002/prot.23202.
Agarwal R., Swaminathan S., and Burley, S. K. Sun . "Structural insight into mechanism and diverse substrate selection strategy of L-ribulokinase". United States. doi:10.1002/prot.23202.
@article{osti_1044005,
title = {Structural insight into mechanism and diverse substrate selection strategy of L-ribulokinase},
author = {Agarwal R. and Swaminathan S. and Burley, S. K.},
abstractNote = {The araBAD operon encodes three different enzymes required for catabolism of L-arabinose, which is one of the most abundant monosaccharides in nature. L-ribulokinase, encoded by the araB gene, catalyzes conversion of L-ribulose to L-ribulose-5-phosphate, the second step in the catabolic pathway. Unlike other kinases, ribulokinase exhibits diversity in substrate selectivity and catalyzes phosphorylation of all four 2-ketopentose sugars with comparable k{sub cat} values. To understand ribulokinase recognition and phosphorylation of a diverse set of substrates, we have determined the X-ray structure of ribulokinase from Bacillus halodurans bound to L-ribulose and investigated its substrate and ATP co-factor binding properties. The polypeptide chain is folded into two domains, one small and the other large, with a deep cleft in between. By analogy with related sugar kinases, we identified {sup 447}{und GG}LPQ{und K}{sup 452} as the ATP-binding motif within the smaller domain. L-ribulose binds in the cleft between the two domains via hydrogen bonds with the side chains of highly conserved Trp126, Lys208, Asp274, and Glu329 and the main chain nitrogen of Ala96. The interaction of L-ribulokinase with L-ribulose reveals versatile structural features that help explain recognition of various 2-ketopentose substrates and competitive inhibition by L-erythrulose. Comparison of our structure to that of the structures of other sugar kinases revealed conformational variations that suggest domain-domain closure movements are responsible for establishing the observed active site environment.},
doi = {10.1002/prot.23202},
journal = {Proteins},
issn = {0887-3585},
number = 1,
volume = 80,
place = {United States},
year = {2012},
month = {1}
}