Crystal Structure and Mutational Studies of Cyanobacterial Branching Enzymes Reveal the Structural Determinants of Reaction Product Specificity
Year: 2025
Type: article
Abstract: ATCC 51142, produce an insoluble branched α-glucan known as cyanobacterial starch. This strain possesses three BE isozymes: cceBE1, cceBE2, and cceBE3. Our previous studies demonstrated that cceBE1 and cceBE2 share similar enzymatic properties and that a "stopper structure" contributes to their preferential production of short chains with a degree of polymerization (DP) of 6 and 7. In contrast, cceBE3 produces small amounts of short (DP5-12) and long (DP30-40) chains and lacks the amino acid sequence corresponding to the stopper structure. To investigate the role of the stopper structure, we constructed a deletion mutant of cceBE1 lacking the stopper structure and characterized its enzymatic properties. The mutant retained catalytic activity but lost the ability to selectively produce glucan chains with DP6 and 7 (transferred chains), providing direct evidence for the stopper structure's role in regulating product chain length. Furthermore, we determined the crystal structure of cceBE3, confirming the absence of the stopper structure. We also identified a unique structural feature in cceBE3, termed subdomain B, located within the predicted substrate-binding site. Deletion of subdomain B led to increased production of short chains (DP3-7), suggesting its involvement in substrate binding and the determination of product specificity. These findings reveal structural determinants of product specificity in cyanobacterial BEs and offer a strategy for engineering BEs to produce novel starch-based materials.
Source: Journal of Applied Glycoscience
DOI: 10.5458/jag.7204105
Authors: Tasuku Tamura, Eiji Suzuki, Ryuichiro Suzuki
Institution: Akita Prefectural University
Language: English
Subfield: Rheumatology
Field: Medicine
Domain: Health Sciences
Open Access status: diamond
OpenAlex ID: W4416377147