title: Costs and benefits of processivity in enzymatic degradation of recalcitrant polysaccharides.
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.
authors.affiliation: Department of Chemistry, Biotechnology, and Food Science, Norwegian University of Life Sciences, P. O. Box 5003, N-1432 As, Norway.
authors.forename: Svein J
authors.initials: SJ
authors.lastname: Horn
authors.forename: Pawel
authors.initials: P
authors.lastname: Sikorski
authors.forename: Jannicke B
authors.initials: JB
authors.lastname: Cederkvist
authors.forename: Gustav
authors.initials: G
authors.lastname: Vaaje-Kolstad
authors.forename: Morten
authors.initials: M
authors.lastname: Sørlie
authors.forename: Bjørnar
authors.initials: B
authors.lastname: Synstad
authors.forename: Gert
authors.initials: G
authors.lastname: Vriend
authors.forename: Kjell M
authors.initials: KM
authors.lastname: Vårum
authors.forename: Vincent G H
authors.initials: VG
authors.lastname: Eijsink
mesh_terms: D002688:Chitinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D011134:Polysaccharides / Q000191:economics / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012706:Serratia marcescens / Q000201:enzymology; D013379:Substrate Specificity
chemical_list: D011134:Polysaccharides; D002688:Chitinases