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1,4-β-Endoglucanase is one of the most important biocatalysts in modern industries. Here, a glycoside hydrolase (GH) family 45 endoglucanase from thermophilic fungus Theilavia terrestris (TtCel45A) was expressed in Pichia pastoris . The recombinant protein shows optimal activity at 60 °C, pH 4–5. The enzyme exhibits extraordinary thermostability that more than 80% activity was detected after heating at 80 °C for 2.5 h. The high resolution crystal structures of apo-form enzyme and that in complex with cellobiose and cellotetraose were solved to 1.36–1.58 Å. The protein folds into two overall regions: one is a six-stranded β-barrel, and the other one consists of several extended loops. Between the two regions lies the substrate-binding channel, which is an open cleft spanning across the protein surface. A continuous substrate-binding cleft from subsite −4 to +3 were clearly identified in the complex structures. Notably, the flexible V–VI loop ( 113 Gly- 114 Gly- 115 Asp- 116 Leu- 117 Gly- 118 Ser) is found to open in the presence of −1 sugar, with D115 and L116 swung away to yield a space to accommodate the catalytic acid D122 and the 2,5 B boat conformation of −1 sugar during transition state. Collectively, we characterized the enzyme properties of P. pastoris -expressed TtCel45A and solved high-resolution crystal structures of the enzyme. These results are of great interests in industrial applications and provide new insights into the fundamental understanding of enzyme catalytic mechanism of GH45 endoglucanases. [ABSTRACT FROM AUTHOR]

S-adenosylhomocysteine (SAH) hydrolase catalyzes the reversible hydrolysis of SAH into adenosine and homocysteine by using NAD + as a cofactor. The enzyme from Thermotoga maritima (tmSAHH) has great potentials in industrial applications because of its hyperthermophilic properties. Here, two crystal structures of tmSAHH in complex with NAD + show both open and closed conformations despite the absence of bound substrate. Each subunit of the tetrameric enzyme is composed of three domains, namely the catalytic domain, the NAD + -binding domain and the C-terminal domain. The NAD + binding mode is clearly observed and a substrate analogue can also be modeled into the active site, where two cysteine residues in mesophilic enzymes are replaced by serine and threonine in tmSAHH. Notably, the C-terminal domain of tmSAHH lacks the second loop region of mesophilic SAHH, which is important in NAD + binding, and thus exposes the bound cofactor to the solvent. The difference explains the higher NAD + requirement of tmSAHH because of the reduced affinity. Furthermore, the feature of missing loop is consistently observed in thermophilic bacterial and archaeal SAHHs, and may be related to their thermostability. [ABSTRACT FROM AUTHOR]