Discovery of a new sialic acid binding region that regulates Siglec-7
Nao Yamakawa, Yu Yasuda, Atsushi Yoshimura, Ami Goshima, Paul R. Crocker, Gérard Vergoten, Yuji Nishiura, Takashi Takahashi, Shinya Hanashima, Kana Matsumoto, Yoshiki Yamaguchi, Hiroshi Tanaka, Ken Kitajima, Chihiro Sato
Abstract
Siglec-7 is a human CD33-like siglec, and is localised predominantly on human natural killer (NK) cells and monocytes. Siglec-7 is considered to function as an immunoreceptor in a sialic acid-dependent manner. However, the underlying mechanisms linking sialic acid-binding and function remain unknown. Here, to gain new insights into the ligand-binding properties of Siglec-7, we carried out in silico analysis and site-directed mutagenesis, and found a new sialic acid-binding region (site 2 containing R67) in addition to the well-known primary ligand-binding region (site 1 containing R124). This was supported by equilibrium dialysis, STD-NMR experiments, and inhibition analysis of GD3-binding toward Siglec-7 using synthetic sialoglycoconjugates and a comprehensive set of ganglioside-based glycoconjugates. Our results suggest that the two ligand-binding sites are potentially controlled by each other due to the flexible conformation of the C-C' loop of Siglec-7.