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Biodegradation of Hemicelluloses
Published in Jean-Luc Wertz, Magali Deleu, Séverine Coppée, Aurore Richel, Hemicelluloses and Lignin in Biorefineries, 2017
Jean-Luc Wertz, Magali Deleu, Séverine Coppée, Aurore Richel
The enzyme is a two-domain, globular protein of 605 amino acid residues and is N-glycosylated at three sites. The first 357 residues constitute an (α / β)8 TIM-barrel domain. The second domain consists of residues 374–559 arranged in a six-stranded β-sandwich, which contains a β-sheet of five parallel β-strands and one antiparallel β-strand, with three α helices on either side of the sheet. A glucose moiety is observed in a pocket at the interface of the two domains, where Asp285 and Glu491 are believed to be involved in catalysis.
Cloning, overexpression, and structural characterization of a novel archaeal thermostable neopullulanase from Desulfurococcus mucosus DSM 2162
Published in Preparative Biochemistry & Biotechnology, 2022
Farzaneh Jafari, Farid Kiani-Ghaleh, Shahrzad Eftekhari, Mehdi Razzaghshoar Razlighi, Nazanin Nazari, Maryam Hajirajabi, Fatima Masoomi Sarvestani, Golnoosh Sharafieh
Investigating the structure and conformation of protein using spectroscopic techniques including Circular Dichroism and X-ray crystallography indicated that generally microbial alpha-amylases have three domains. Domain A, the central domain of the enzyme, has the central structure of the TIM barrel with Asp 231, Glu 261 and Asp 328 residues at the active site. On the other hand, the domains B and C are located on the sides of the barrel. Domain B is located between the third strand and the third helix of the barrel and has created a pseudo-beta structure which is possibly responsible for the difference in substrate specificity and alpha-amylase stability.[44] The C-terminal of C-domain contains the Greek key motif which seems to be effective in transferring the enzyme across the outer membrane, binding the enzyme to starch, and its thermal stability. Further, some amylases have a carbohydrate-binding module (CBM) for hydrolysis of the insoluble starch. The thermostability of a protein could be due to several synergistic effects, particularly, the presence of the α-helix structures, since breaking the regular structure during denaturation requires a great deal of energy.[45] Non-catalytic CBM domain consists of 40 to 200 individual amino acids, which facilitates the binding of enzymes to a substrate, thus enhancing hydrolysis. Approximately 10% of amylolytic enzymes have separate chemically bound starch binding sites. For example, in Lactobacilli, these dominant domains have been found in three amylase sequences with the sequence similarity being 96%. In the present analysis, the far UV-CD spectrum of DSMA (Figure 4) shows a positive peak at 192 nm and two negative peaks at 208 and 222 nm, which are typical of α-helical protein.[30,46]. Meanwhile, the percentage of stable folding pattern in a protein could be estimated according to the analysis of DichroWeb software. Based on the results, DSMA was estimated to have 47.3% of α-helix and 31.6% of β-sheet structure, whereas 8.5% of its structure was turn and 12.6% was random coil.