Determination of beta-glucosidase aggregating factor (BGAF) binding and polymerization regions on the maize beta-glucosidase isozyme Glu1.
Beta-glucosidases (Glu1 and Glu2) in maize specifically interact with a lectin called beta-glucosidase aggregating factor (BGAF). We have shown that the N-terminal (Glu(50)-Val(145)) and the C-terminal (Phe(466)-Ala(512)) regions of maize Glu1 are involved in binding to BGAF. Sequence comparison between sorghum beta-glucosidases (dhurrinases, which do not bind to BGAF) and maize beta-glucosidases, and the 3D-structure of Glu1 suggested that the BGAF-binding site on Glu1 is much smaller than predicted previously. To define more precisely the BGAF-binding site, we constructed additional chimeric beta-glucosidases. The results showed that a region spanning 11 amino acids (Ile(72)-Thr(82)) on Glu1 is essential and sufficient for BGAF binding, whereas the extreme N-terminal region Ser(1)-Thr(29), together with C-terminal region Phe(466)-Ala(512), affects the size of Glu1-BGAF complexes. The dissociation constants (K(d)) of chimeric beta-glucosidase-BGAF interactions also demonstrated that the extreme N-terminal and C-terminal regions are important but not essential for binding. To confirm the importance of Ile(72)-Thr(82) on Glu1 for BGAF binding, we constructed a chimeric sorghum beta-glucosidase, Dhr2 (C-11, Dhr2 whose Val(72)-Glu(82) region was replaced with the Ile(72)-Thr(82) region of Glu1). C-11 binds to BGAF, indicating that the Ile(72)-Thr(82) region is indeed a major interaction site on Glu1 involved in BGAF binding.
Yu
Hyun Young
HY
Department of Biological Sciences, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061-0406, USA. yhy0922@vt.edu
Kittur
Farooqahmed S
FS
Bevan
David R
DR
Esen
Asim
A
D000595:Amino Acid Sequence; D001665:Binding Sites; D044602:Cellulases / Q000737:chemistry*; D007527:Isoenzymes; D037102:Lectins / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D045868:Sorghum / Q000201:enzymology / Q000235:genetics; D003313:Zea mays / Q000737:chemistry / Q000201:enzymology*
D007527:Isoenzymes; D037102:Lectins; C545968:beta-glucosidase aggregating factor; D044602:Cellulases