tailieunhanh - Báo cáo khoa học: Substrate specificity and catalytic mechanism of a xyloglucan xyloglucosyl transferase HvXET6 from barley (Hordeum vulgare L.)

A family 16 glycoside hydrolase, xyloglucan xyloglucosyl transferase (EC ), also known as xyloglucan endotransglycosylase (XET), and designated isoenzyme HvXET6, was purified approximately 400-fold from extracts of young barley seedlings. | Substrate specificity and catalytic mechanism of a xyloglucan xyloglucosyl transferase HvXET6 from barley Hordeum vulgare L. Maria Hrmova1 Vladimir Farkas2 Andrew J. Harvey1 Jelle Lahnstein1 Bente Wischmann3 Nomchit Kaewthai4 Ines Ezcurra4 Tuula T. Teeri4 and Geoffrey B. Fincher1 1 Schoolof Agriculture Food and Wine Australian Centre for Plant FunctionalGenomics University of Adelaide Australia 2 Institute of Chemistry Centre of Excellence GLYCOBIOS Slovak Academy of Sciences Bratislava Slovak Republic 3 Novozymes A S Bagsvaerd Denmark 4 Schoolof Biotechnology RoyalInstitute of Biotechnology AlbaNova University Centre Stockholm Sweden Keywords glycoside hydrolase family 16 molecular modelling phylogenetic analyses plant cell walls reaction mechanisms Correspondence M. Hrmova and G. Fincher Schoolof Agriculture Food and Wine Australian Centre for Plant Functional Genomics University of Adelaide Waite Campus Glen Osmond SA 5064 Australia Fax 61 8 830 37102 Tel 61 8 830 37280 37296 E-mail Received 8 August 2008 revised 2 October 2008 accepted 10 November 2008 doi A family 16 glycoside hydrolase xyloglucan xyloglucosyl transferase EC also known as xyloglucan endotransglycosylase XET and designated isoenzyme HvXET6 was purified approximately 400-fold from extracts of young barley seedlings. The complete amino acid sequence of HvXET6 was deduced from the nucleotide sequence of a near full-length cDNA in combination with tryptic peptide mapping. An additional five to six isoforms or post-translationally modified XET enzymes were detected in crude seedling extracts of barley. The HvXET6 isoenzyme was expressed in Pichia pastoris characterized and compared with the previously purified native HvXET5 isoform. Barley HvXET6 has a similar apparent molecular mass of 33-35 kDa to the previously purified HvXET5 isoenzyme but the two isoenzymes differ in their isoelectric points pH