Mutation of a conserved tryptophan residue in the CBM3c of a GH9 endoglucanase inhibits activity

Su Jung Kim, So Hyeong Kim, Sang Kyu Shin, Jeong Eun Hyeon, Sung Ok Han

Research output: Contribution to journalArticlepeer-review

8 Citations (Scopus)


The presence of the family of 3c cellulose binding module (CBM3c) is important for the catalytic activity of family 9 endoglucanases such as the EngZ from Clostridium cellulovorans. To determine the role of CBM3c in catalytic activity, we made a tryptophan to alanine substitution because tryptophan can bind strongly to both substrates and other amino acids. The conserved tryptophan substitution (W483A) did not influence substrate binding, but it reduced enzyme activity to 10–14% on both amorphous and crystalline cellulose. CBM3c is directly involved in the endoglucanase reaction independent of substrate binding. EngZ W483A was also inactivated independent of substrate concentrations. Specially, EngZ W483A restored its catalytic base activity (31.6 ± 1.2 U/nM) which is similar to the wild-type (29.4 ± 0.3 U/nM) on Avicel in the presence of 50 mM sodium azide which is instead of catalytic base reaction. These results suggest that CBM3c is deeply involved in the cellulolytic reaction, specifically at the catalytic base region. Moreover, EngZ W483A was also easily denatured by DTT, an outer disulfide bond breaker, compared to the wild-type. CBM3c could influence the surface stability. These features of CBM3c result from the hydrophobic interaction of tryptophan with the catalytic domain that is unrelated to substrate binding.

Original languageEnglish
Pages (from-to)159-166
Number of pages8
JournalInternational Journal of Biological Macromolecules
Publication statusPublished - 2016 Nov 1


  • CD and CBM interaction
  • Catalytic base
  • Cellulose binding module 3c
  • Disulfide bond
  • Family 9 endoglucanase
  • Initial cellulolytic reaction

ASJC Scopus subject areas

  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Economics and Econometrics
  • Energy(all)


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