Cloning, sequencing, and expression of the tulip bulb chitinase-1 cdna

Takeshi Yamagami, Kazuki Tsutsumi, Masatsune Ishiguro

研究成果: ジャーナルへの寄稿学術誌査読

3 被引用数 (Scopus)

抄録

A cDNA encoding tulip bulb chitinase-1 (TBC-1) was cloned using a combination of immunoscreening from a λ ZAP cDNA library with anti-TBC-1 antiserum and the 5′ rapid amplification of cDNA end (RACE) method, and sequenced. The cDNA consists of 1,106 nucleotides and included an open reading frame encoding a polypeptide of 314 amino acids. Comparison of the deduced amino acid sequence and the determined protein sequence indicated the presence of a signal peptide and an extra peptide composed of 26 and 13 amino acids at the N- and C-termini, respectively. The deduced sequence of TBC-1 had 10-20% and 63% sequence similarities to plant class III chitinases and gladiolus bulb class IIIb chitinase (GBC-a), respectively. The cDNA encoding mature TBC-1 was amplified by polymerase chain reaction (PCR), ligated into the expression vector pET-22b, and expressed in Escherichia coli BL21(DE3). The recombinant TBC-1 (rTBC-1) expressed in E. coli was purified by gel filtration followed by ion-exchange chromatography. Specific activity of the rTBC-1 was almost same as the authentic TBC-1 toward glycolchitin. This is the first report on the cDNA cloning of a class III chitinase having C-terminal extra peptide.

本文言語英語
ページ(範囲)1394-1401
ページ数8
ジャーナルBioscience, Biotechnology and Biochemistry
64
7
DOI
出版ステータス出版済み - 2000

!!!All Science Journal Classification (ASJC) codes

  • バイオテクノロジー
  • 分析化学
  • 生化学
  • 応用微生物学とバイオテクノロジー
  • 分子生物学
  • 有機化学

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