Crystal Structure of α-Galactosidase from Thermus thermophilus : Insight into Hexamer Assembly and Substrate Specificity

• Published in: Journal of agricultural and food chemistry Vol. 68; no. 22; pp. 6161 - 6169
• Main Authors: Chen, Sheng-Chia, Wu, Szu-Pei, Chang, Yu-Yung, Hwang, Tzann-Shun, Lee, Tzong-Huei, Hsu, Chun-Hua
• Format: Journal Article
• Language: English
• Published: United States 03.06.2020
• Subjects: alpha-Galactosidase - chemistry; alpha-Galactosidase - genetics; alpha-Galactosidase - metabolism; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Biocatalysis; Catalytic Domain; Crystallography, X-Ray; Galactose - metabolism; Protein Domains; Protein Multimerization; Substrate Specificity; Thermus thermophilus - chemistry; Thermus thermophilus - enzymology; Thermus thermophilus - genetics; Thermus thermophilus - metabolism; stachyose; thermostable; α-galactosidase; substrate specificity; hexamer assembly
• ISSN: 0021-8561; 1520-5118
• DOI: 10.1021/acs.jafc.0c00871

α-Galactosidase catalyzes the hydrolysis of a terminal α-galactose residue in galacto-oligosaccharides and has potential in various industrial applications and food processing. We determined the crystal structures of α-galactosidase from the thermophilic microorganism (TtGalA) and its complexes with pNPGal and stachyose. The monomer folds into an N-terminal domain, a catalytic (β/α) barrel domain, and a C-terminal domain. The domain organization is similar to that of the other family of 36 α-galactosidases, but TtGalA presents a cagelike hexamer. Structural analysis shows that oligomerization may be a key factor for the thermal adaption of TtGalA. The structure of TtGalA complexed with stachyose reveals only the existence of one -1 subsite and one +1 subsite in the active site. Structural comparison of the stachyose-bound complexes of TtGalA and GsAgaA, a tetrameric enzyme with four subsites, suggests evolutionary divergence of substrate specificity within the GH36 family of α-galactosidases. To the best of our knowledge, the crystal structure of TtGalA is the first report of a quaternary structure as a hexameric assembly in the α-galactosidase family.