Cyanobacterial branching enzymes bind to α-glucan via surface binding sites.

Yousra El Mannai, Ryota Deto, Miho Kuroki, Ryuichiro Suzuki, Eiji Suzuki

Archives of biochemistry and biophysics 2021 May 15

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Besides their catalysis, specific interactions between starch/glycogen processing enzymes and their substrates have been reported. Multiple branching enzyme (BE) isoforms, BE1, BE2, and BE3, have been found in a limited number of cyanobacterial species that are characterized by amylopectin accumulation. Seven surface binding sites (SBSs) located away from the active site have been identified in crystal structures of cyanobacterial BE1 from Crocosphaera subtropica (Cyanothece sp.) ATCC 51142 (51142BE1). In the present study, binding affinity toward amylopectin, amylose, and glycogen was investigated for wild-type 51142BE1 and its mutants (residues at SBSs important for sugar-binding were replaced by alanine). These enzymes showed retarded mobility during electrophoresis in non-denaturing polyacrylamide gels in the presence of polysaccharides. This was caused by interactions between the enzymes and the polysaccharides, enabling calculation of the dissociation constants (Kd values) of the enzymes toward the polysaccharides. Mutational analysis indicated that particular domains of the protein (domains A and C) were involved in the polysaccharide binding. Kd values toward the polysaccharides were also measured for 10 BE isoforms (five BE1, three BE2, and two BE3) from 5 cyanobacterial strains. All BEs displayed much lower Kd values (higher affinity) toward amylopectin and amylose than toward glycogen, as described for plant BEs. In addition, one BE2 displayed exceptionally high Kd values (low affinity), while two BE3 exhibited multiple Kd values to all polysaccharides. These results could be ascribed to sequence variations in the SBSs, irrespective of the catalytic specificity. Copyright © 2021 Elsevier Inc. All rights reserved.

Citation

Yousra El Mannai, Ryota Deto, Miho Kuroki, Ryuichiro Suzuki, Eiji Suzuki. Cyanobacterial branching enzymes bind to α-glucan via surface binding sites. Archives of biochemistry and biophysics. 2021 May 15;702:108821