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Trp17 and Glu20 Residues in Conserved WMN(D/E)PN Motif Are Essential for Aspergillus ficuum Endoinulinase (EC Activity

Seockyu Park, Yeonsoo Han, Heeun Kim, Songyi Song, Tai-Boong Uhm, and Keon-Sang Chae*

Division of Biological Sciences and Research Center for Industrial Development of Biofood Materials, Chonbuk National University, Chonju, Chonbuk 561-756, Republic of Korea; fax: +82 63 270 3345; E-mail: chaeks@moak.chonbuk.ac.kr

* To whom correspondence should be addressed.

Received November 22, 2002; Revision received December 3, 2002
The importance of the WMN(D/E)PN motif, which is well conserved among beta-fructofuranosidases grouped in the glycosylhydrolase family 32, in Aspergillus ficuum endoinulinase was accessed. Each mutant enzyme generated by site-directed mutagenesis of Trp17 in the conserved motif to Gln, Leu, Ser, Pro, Thr, or Met had an activity of less than 1% of the wild type. Another mutant enzyme obtained by mutation of Glu20 in the motif to Ser, Leu, Thr, Gln, Ala, or Val had an enzyme activity of less than 1% of the wild type. Furthermore, the E20D mutant enzyme, in which Glu20 in the conserved motif was replaced with Asp, had 1.1% of the wild type activity. These results clearly indicated that Trp17 and Glu20 are essential for the enzyme activity.
KEY WORDS: Aspergillus ficuum, endoinulinase, glutamic acid, inuB, tryptophan