Reduced type-A carbohydrate-binding module interactions to cellulose I leads to improved endocellulase activity

Bhargava Nemmaru, Nicholas Ramirez, Cindy J. Farino, John M. Yarbrough, Nicholas Kravchenko, Shishir P.S. Chundawat

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

Abstract

Dissociation of nonproductively bound cellulolytic enzymes from cellulose is hypothesized to be a key rate-limiting factor impeding cost-effective biomass conversion to fermentable sugars. However, the role of carbohydrate-binding modules (CBMs) in enabling nonproductive enzyme binding is not well understood. Here, we examine the subtle interplay of CBM binding and cellulose hydrolysis activity for three models type-A CBMs (Families 1, 3a, and 64) tethered to multifunctional endoglucanase (CelE) on two distinct cellulose allomorphs (i.e., cellulose I and III). We generated a small library of mutant CBMs with varying cellulose affinity, as determined by equilibrium binding assays, followed by monitoring cellulose hydrolysis activity of CelE–CBM fusion constructs. Finally, kinetic binding assays using quartz crystal microbalance with dissipation were employed to measure CBM adsorption and desorption rate constants (Formula presented.) and (Formula presented.), respectively, towards nanocrystalline cellulose derived from both allomorphs. Overall, our results indicate that reduced CBM equilibrium binding affinity towards cellulose I alone, resulting from increased desorption rates ((Formula presented.)) and reduced effective adsorption rates ((Formula presented.)), is correlated to overall improved endocellulase activity. Future studies could employ similar approaches to unravel the role of CBMs in nonproductive enzyme binding and develop improved cellulolytic enzymes for industrial applications.

Original languageEnglish (US)
Pages (from-to)1141-1151
Number of pages11
JournalBiotechnology and Bioengineering
Volume118
Issue number3
DOIs
StatePublished - Mar 2021

All Science Journal Classification (ASJC) codes

  • Applied Microbiology and Biotechnology
  • Bioengineering
  • Biotechnology

Keywords

  • carbohydrate-binding module
  • cellulose III
  • endocellulases
  • nonproductive binding
  • protein adsorption
  • quartz crystal microbalance with dissipation

Fingerprint Dive into the research topics of 'Reduced type-A carbohydrate-binding module interactions to cellulose I leads to improved endocellulase activity'. Together they form a unique fingerprint.

Cite this