caa a22 4500 605501831 CHVBK 20210128100602.0 cr unu---uuuuu 210128e20150201xx s 000 0 eng 10.1007/s00253-014-6269-3 doi (NATIONALLICENCE)springer-10.1007/s00253-014-6269-3 Insights into the substrate specificity and synergy with mannanase of family 27 α-galactosidases from Neosartorya fischeri P1 [Elektronische Daten] [Huimin Wang, Rui Ma, Pengjun Shi, Huoqing Huang, Peilong Yang, Yaru Wang, Yunliu Fan, Bin Yao] Thermophilic Neosartorya fischeri P1 is an excellent carbohydrate-active enzyme (CAZyme) producer. Two α-galactosidases of GH (glycoside hydrolase ) family 27 with a very low sequence identity (28.7%), Gal27A and Gal27B, were identified in strain P1 and functionally expressed in Pichia pastoris. In comparison to other characterized GH27 fungal counterparts, rGal27B has a higher temperature optimum (75°C) and better thermostability (>50% activity at 70°C for 15min), and rGal27A shows stability over the broadest pH range (pH2.0-12.0). Moreover, great distinctions lie in the two enzymes. When using pNPG as the substrate, rGal27B had a higher turnover number (1621.4 vs. 368.3s−1) but lower affinity (2.84 vs. 0.8mM) and catalytic efficiency (460.8 vs. 580.3s−1mM−1) than rGal27A. rGal27B acted on galacto-oligosaccharides, whereas rGal27A was active on polymeric substrates. Although both enzymes showed synergy in galactomannan degradation when combined with a β-mannanase of the same strain, enzyme combinations including rGal27A released more reducing sugars (up to 11.67-fold). Homology modeling predicts different loops in N. fischeri α-galactosidases, highlighting the larger tunnel structure in Gal27A to accommodate/bind branched galactomannan with high galactose contents. Phylogenetic analysis reveals the far relationship of Gal27A and Gal27B that they may evolve in different action modes, and their coexistence widens the substrate spectrum for nutrient utilization. This study illustrates the substrate profiles and synergistic mechanism of GH27 α-galactosidases of different structures. Springer-Verlag Berlin Heidelberg, 2014 Neosartorya fischeri P1 nationallicence α-Galactosidase nationallicence Glycoside hydrolase (GH) 27 nationallicence Substrate and product profile nationallicence Synergy nationallicence Wang Huimin Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut Ma Rui Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut Shi Pengjun Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut Huang Huoqing Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut Yang Peilong Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut Wang Yaru Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut Fan Yunliu Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, 100081, Beijing, People's Republic of China aut Yao Bin Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/3(2015-02-01), 1261-1272 0175-7598 99:3<1261 2015 99 253 https://doi.org/10.1007/s00253-014-6269-3 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-014-6269-3 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Wang Huimin Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut NATIONALLICENCE 700 1- Ma Rui Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut NATIONALLICENCE 700 1- Shi Pengjun Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut NATIONALLICENCE 700 1- Huang Huoqing Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut NATIONALLICENCE 700 1- Yang Peilong Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut NATIONALLICENCE 700 1- Wang Yaru Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut NATIONALLICENCE 700 1- Fan Yunliu Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, 100081, Beijing, People's Republic of China aut NATIONALLICENCE 700 1- Yao Bin Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/3(2015-02-01), 1261-1272 0175-7598 99:3<1261 2015 99 253