Capacity of thermomonospora alba Xy1A to impart thermostability in family F/10 chimeric xylanases

Mohammad Mainul Ahsan, Satoshi Kaneko, Qin Wang, Kei Yura, Mitiko Go, Kiyoshi Hayash*


研究成果: Article査読

15 被引用数 (Scopus)


To reveal structure-function relationships of family F/10 glycanases, an in vitro molecular level shuffling experiment was conducted to accumulate useful amino acid residues from two homologous F/10 xylanases, FXYN of Streptomyces olivaceoviridis E-86 and Xy1A of Thermomonospora alba ULJB1, into a single chimeric xylanase. The parent genes were shuffled by crossovers at selected module borders using self-priming Polymerase Chain Reaction (PCR)s. The shuffled constructs, designated as FXYN-M3/4-Xy1A, FXYN-M9/10-Xy1A, and FXYN-M14/15-Xy1A were cloned and their nucleotide sequences were confirmed. Two chimera, FXYN-M3/4-Xy1A and FXYN-M14/15-Xy1A, demonstrated activity against RBB-xylan and were over-expressed as His-tag fusion proteins under control of T5 promoter of pQE60. The homogeneously pure chimeric proteins, FXYN-M3/4-Xy1A and FXYN-M14/15-Xy1A showed improved thermal and pH profiles compared to those of one of the parents, FXYN. This was apparently due to the influence of amino acids inherited from thermophilic Xy1A. Measured K(m) and kcat values were closer to those of the other parent, Xy1A. Interestingly, a significant level of heat tolerance up to 60°C, was recorded for FXYN-M3/4-Xy1A in comparison to only 40°C for FXYN-M14/15-Xy1A though their temperature optima did not correlates with their thermal stability. These results indicated that the amino acid residues of the larger T. alba Xy1A DNA fragment present in FXYN-M3/4-Xy1A were responsible for inducing its thermal stability. (C) 2001 Elsevier Science Inc.

ジャーナルEnzyme and Microbial Technology
出版ステータスPublished - 2001 1月 2

ASJC Scopus subject areas

  • バイオテクノロジー
  • バイオエンジニアリング
  • 生化学
  • 応用微生物学とバイオテクノロジー


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