TY  - JOUR
A1  - Vaaje-Kolstad, G.
A1  - Vasella, A.
A1  - Peter, Martin G.
A1  - Netter, C.
A1  - Houston, Douglas R.
A1  - Westereng, B.
A1  - Synstad, Bjoenar
A1  - Eijsink, Vincent G. H.
A1  - van Aalten, Daan M. F.
T1  - Interactions of a family 18 chitinase with the designed inhibitor HM508 and its degradation product, chitobiono- delta-lactone
N2  - We describe enzymological and structural analyses of the interaction between the family 18 chitinase ChiB from Serratia marcescens and the designed inhibitor N,N'-diacetylchitobionoxime-N-phenylcarbamate (HM508). HM508 acts as a competitive inhibitor of this enzyme with a K-i in the 50 muM range. Active site mutants of ChiB show K-i values ranging from 1 to 200 muM, providing insight into some of the interactions that determine inhibitor affinity. Interestingly, the wild type enzyme slowly degrades HM508, but the inhibitor is essentially stable in the presence of the moderately active D142N mutant of ChiB. The crystal structure of the D142N-HM508 complex revealed that the two sugar moieties bind to the -2 and -1 subsites, whereas the phenyl group interacts with aromatic side chains that line the +1 and +2 subsites. Enzymatic degradation of HM508, as well as a Trp-->Ala mutation in the +2 subsite of ChiB, led to reduced affinity for the inhibitor, showing that interactions between the phenyl group and the enzyme contribute to binding. Interestingly, a complex of enzymatically degraded HM508 with the wild type enzyme showed a chitobiono-delta- lactone bound in the -2 and -1 subsites, despite the fact that the equilibrium between the lactone and the hydroxy acid forms in solution lies far toward the latter. This shows that the active site preferentially binds the E-4 conformation of the -1 sugar, which resembles the proposed transition state of the reaction
Y1  - 2004
SN  - 0021-9258
ER  - 
TY  - JOUR
A1  - Houston, Douglas R.
A1  - Shiomi, Kazuro
A1  - Arai, Noriko
A1  - Omura, Satoshi
A1  - Peter, Martin G.
A1  - Turberg, Andreas
A1  - Synstad, Bjoenar
A1  - Eijsink, Vincent G. H.
A1  - Van Aalten, Daan M. F.
T1  - High-resolution structures of a chitinase complexed with natural product cyclopentapeptide inhibitors : mimicry of carbohydrate substrate
N2  - Over the past years, family 18 chitinases have been validated as potential targets for the design of drugs against human pathogens that contain or interact with chitin during their normal life cycles. Thus far, only one potent chitinase inhibitor has been described in detail, the pseudotrisaccharide allosamidin. Recently, however, two potent natural-product cyclopentapeptide chitinase inhibitors, argifin and argadin, were reported. Here, we describe high- resoln. crystal structures that reveal the details of the interactions of these cyclopeptides with a family 18 chitinase. The structures are examples of complexes of a carbohydrate-processing enzyme with high-affinity peptide-based inhibitors and show in detail how the peptide backbone and side chains mimic the interactions of the enzyme with chitooligosaccharides. Together with enzymol. characterization, the structures explain why argadin shows an order of magnitude stronger inhibition than allosamidin, whereas argifin shows weaker inhibition. The peptides bind to the chitinase in remarkably different ways, which may explain the differences in inhibition consts. The two complexes provide a basis for structure-based design of potent chitinase inhibitors, accessible by std. peptide chem.
Y1  - 2002
ER  - 
TY  - JOUR
A1  - Eijsink, Vincent G. H.
A1  - Synstad, Bjoenar
A1  - Gaseidnes, Sigrid
A1  - Komander, David
A1  - Houston, Douglas R.
A1  - Peter, Martin G.
A1  - van Aalten, Daan M. F.
T1  - Structure and function of chitinolytic enzymes
N2  - The recent work on a variety of family 18 chitonolytic enzymes has yielded important data concerning the structure, substrate-binding, catalysis, inhibitor-binding and even dynamics. These data have been useful in helping to better understand the roles of various types of chitinases in chitin hydrolysis, to rationally engineer the properties of these enzymes, thus making them more suitable as biocatalysts, and to study and understand the effectiveness of natural and designed chitinase inhibitors, which may be of medical interest. On the other hand, the recent work on ChiB shows that catalysis in family 18 chitinases is a highly complicated process, involving larger parts of the enzyme and dynamics. Thus, despite recent discoveries, there is still a lot more to discover about how these enzyme work.
Y1  - 2003
SN  - 82-471-5901-5
ER  -