Costs and benefits of processivity in enzymatic degradation of recalcitrant polysaccharides

doi:10.1073/pnas.0608909103

Costs and benefits of processivity in enzymatic degradation of recalcitrant polysaccharides

*Department of Chemistry, Biotechnology, and Food Science, Norwegian University of Life Sciences, P. O. Box 5003, N-1432 Ås, Norway;
^†Department of Physics and
^§Norwegian Biopolymer Laboratory (NOBIPOL), Department of Biotechnology, Norwegian University of Science and Technology, 7491 Trondheim, Norway; and
^‡Center for Molecular and Biomolecular Informatics, Nijmegen Center for Molecular Life Sciences, Radboud University, 6525 ED, Nijmegen, The Netherlands

Communicated by T. Kent Kirk, University of Wisconsin, Madison, WI, October 9, 2006 (received for review July 9, 2006)

Abstract

Many enzymes that hydrolyze insoluble crystalline polysaccharides such as cellulose and chitin guide detached single-polymer chains through long and deep active-site clefts, leading to processive (stepwise) degradation of the polysaccharide. We have studied the links between enzyme efficiency and processivity by analyzing the effects of mutating aromatic residues in the substrate-binding groove of a processive chitobiohydrolase, chitinase B from Serratia marcescens. Mutation of two tryptophan residues (Trp-97 and Trp-220) close to the catalytic center (subsites +1 and +2) led to reduced processivity and a reduced ability to degrade crystalline chitin, suggesting that these two properties are linked. Most remarkably, the loss of processivity in the W97A mutant was accompanied by a 29-fold increase in the degradation rate for single-polymer chains as present in the soluble chitin-derivative chitosan. The properties of the W220A mutant showed a similar trend, although mutational effects were less dramatic. Processivity is thought to contribute to the degradation of crystalline polysaccharides because detached single-polymer chains are kept from reassociating with the solid material. The present results show that this processivity comes at a large cost in terms of enzyme speed. Thus, in some cases, it might be better to focus strategies for enzymatic depolymerization of polysaccharide biomass on improving substrate accessibility for nonprocessive enzymes rather than on improving the properties of processive enzymes.

Footnotes

^¶To whom correspondence should be addressed. E-mail: vincent.eijsink{at}umb.no
Author contributions: S.J.H., P.S., G.V., K.M.V., and V.G.H.E. designed research; S.J.H., P.S., J.B.C., G.V.-K., M.S., B.S., and K.M.V. performed research; P.S. and K.M.V. contributed new reagents/analytic tools; S.J.H., P.S., G.V.-K., M.S., B.S., G.V., K.M.V., and V.G.H.E. analyzed data; and S.J.H., G.V.-K., and V.G.H.E. wrote the paper.
The authors declare no conflict of interest.
Abbreviations:

CBM,

carbohydrate-binding module;

ChiB,

chitinase B from Serratia marcescens;

DP,

degree of polymerization