Heterogonous expression and characterization of a plant class IV chitinase from the pitcher of the carnivorous plant Nepenthes alata

Kana Ishisaki, Yuji Honda, Hajime Taniguchi, Naoya Hatano, Tatsuro Hamada

Research output: Contribution to journalArticlepeer-review

25 Citations (Scopus)

Abstract

A class IV chitinase belonging to the glycoside hydrolase 19 family from Nepenthes alata (NaCHIT1) was expressed in Escherichia coli. The enzyme exhibited weak activity toward polymeric substrates and significant activity toward (GlcNAc) n [-1,4-linked oligosaccharide of GlcNAc with a polymerization degree of n (n = 4-6)]. The enzyme hydrolyzed the third and fourth glycosidic linkages from the non-reducing end of (GlcNAc) 6. The pH optimum of the enzymatic reaction was 5.5 at 37°C. The optimal temperature for activity was 60°C in 50 mM sodium acetate buffer (pH 5.5). The anomeric form of the products indicated that it was an inverting enzyme. The k cat/K m of the (GlcNAc) n hydrolysis increased with an increase in the degree of polymerization. Amino acid sequence alignment analysis between NaCHIT1 and a class IV chitinase from a Picea abies (Norway spruce) suggested that the deletion of four loops likely led the enzyme to optimize the (GlcNAc) n hydrolytic reaction rather than the hydrolysis of polymeric substrates.

Original languageEnglish
Pages (from-to)345-351
Number of pages7
JournalGlycobiology
Volume22
Issue number3
DOIs
Publication statusPublished - Mar 2012
Externally publishedYes

Keywords

  • GH family 19
  • Nepenthes
  • carnivorous plant
  • chitinase
  • class IV

ASJC Scopus subject areas

  • Biochemistry

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