Dimension, shape, and conformational flexibility of a two domain fungal cellulase in solution probed by small angle X-ray scattering

J Biol Chem. 2002 Oct 25;277(43):40887-92. doi: 10.1074/jbc.M205404200. Epub 2002 Aug 16.

Abstract

Cellulase Cel45 from Humicola insolens has a modular structure with a catalytic module and a cellulose-binding module (CBM) separated by a 36 amino acid, glycosylated, linker peptide. The solution conformation of the entire two domain Cel45 protein as well as the effect of the length and flexibility of the linker on the spatial arrangement of the constitutive modules were studied by small angle x-ray scattering combined with the known three-dimensional structure of the individual modules. The measured dimensions of the enzyme show that the linker exhibits an extended conformation leading to a maximum extension between the two centers of mass of each module corresponding to about four cellobiose units on a cellulose chain. The glycosylation of the linker is the key factor defining its extended conformation, and a five proline stretch mutation on the linker was found to confer a higher rigidity to the enzyme. Our study shows that the respective positioning of the catalytic module and the CBM onto the insoluble substrate is most likely influenced by the linker structure and flexibility. Our results are consistent with a model where cellulases can move on the surface of cellulose with a caterpillar-like displacement with free energy restrictions.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Ascomycota / enzymology*
  • Cellulase / chemistry*
  • Glycosylation
  • Mitosporic Fungi / enzymology*
  • Models, Molecular
  • Protein Conformation
  • Scattering, Radiation
  • X-Rays

Substances

  • Cellulase