Kinetic Characterization of 100 Glycoside Hydrolase Mutants Enables the Discovery of Structural Features Correlated with Kinetic Constants


Abstract

The use of computational modeling algorithms to guide the design of novel enzyme catalysts is a rapidly growing field. Force-field based methods have now been used to engineer both enzyme specificity and activity. However, the proportion of designed mutants with the intended function is often less than ten percent. One potential reason for this is that current force-field based approaches are trained on indirect measures of function rather than direct correlation to experimentally-determined functional effects of mutations. We hypothesize that this is partially due to the lack of data sets for which a large panel of enzyme variants has been produced, purified, and kinetically characterized. Here we report the kcat and KM values of 100 purified mutants of a glycoside hydrolase enzyme. We demonstrate the utility of this data set by using machine learning to train a new algorithm that enables prediction of each kinetic parameter based on readily-modeled structural features. The generated dataset and analyses carried out in this study not only provide insight into how this enzyme functions, they also provide a clear path forward for the improvement of computational enzyme redesign algorithms.

Submission Details

ID: 8yraQWia3

Submitter: Barry Olafson

Submission Date: Nov. 21, 2016, 2:51 p.m.

Version: 1

Publication Details
Carlin DA;Caster RW;Wang X;Betzenderfer SA;Chen CX;Duong VM;Ryklansky CV;Alpekin A;Beaumont N;Kapoor H;Kim N;Mohabbot H;Pang B;Teel R;Whithaus L;Tagkopoulos I;Siegel JB,PLoS One (2016) Kinetic Characterization of 100 Glycoside Hydrolase Mutants Enables the Discovery of Structural Features Correlated with Kinetic Constants. PMID:26815142
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)