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To examine the effect of a conformational constraint introduced into the Arg-Gly-Asp (RGD) sequence on cell adhesion activity, we constructed a mutant protein by inserting an RGD-containing sequence flanked by two Cys residues between Val74 and Asn75 of human lysozyme. The CRGDSC-inserted lysozyme was expressed in yeast, purified, and designated as Cys-RGD4. Using baby hamster kidney cells, Cys-RGD4 was shown to possess even higher cell adhesion activity than that of the RGDS-inserted lysozyme, RGD4. The Cys-RGD4 protein was co-crystallized with a lysozyme inhibitor, tri-N-acetylchitotriose, and the three-dimensional structure was determined at 1.6-A resolution by x-ray crystallography. In contrast to RGD4, the inserted RGD-containing region of Cys-RGD4 was well defined. The structural analysis revealed that the two inserted Cys residues form a new disulfide bond in Cys-RGD4, as expected, and that the RGD region assumes a type II' beta-turn conformation of Gly-Asp with a hydrogen bond between the C = O of Arg and the H-N of Ser. In addition, it was confirmed that two more hydrogen bonds are present in the RGD region of the Cys-RGD4 lysozyme. These results suggest that the conformation of the RGD-containing region is rigid and stable in the Cys-RGD4 molecule and that the type II' beta-turn structure of RGD is essential for binding to integrins with high affinity.
T Yamada, and
H Song, and
K Inaka, and
Y Shimada, and
M Kikuchi, and
M Matsushima
October 1994,
Biochemistry,
T Yamada, and
H Song, and
K Inaka, and
Y Shimada, and
M Kikuchi, and
M Matsushima
March 1999,
European journal of biochemistry,
T Yamada, and
H Song, and
K Inaka, and
Y Shimada, and
M Kikuchi, and
M Matsushima
May 1993,
The Journal of biological chemistry,
T Yamada, and
H Song, and
K Inaka, and
Y Shimada, and
M Kikuchi, and
M Matsushima
April 1995,
Molecular immunology,
T Yamada, and
H Song, and
K Inaka, and
Y Shimada, and
M Kikuchi, and
M Matsushima
January 1997,
Tanpakushitsu kakusan koso. Protein, nucleic acid, enzyme,
T Yamada, and
H Song, and
K Inaka, and
Y Shimada, and
M Kikuchi, and
M Matsushima
August 1997,
Blood cells, molecules & diseases,
T Yamada, and
H Song, and
K Inaka, and
Y Shimada, and
M Kikuchi, and
M Matsushima
August 1985,
The Journal of experimental medicine,
T Yamada, and
H Song, and
K Inaka, and
Y Shimada, and
M Kikuchi, and
M Matsushima
October 1995,
FEBS letters,
T Yamada, and
H Song, and
K Inaka, and
Y Shimada, and
M Kikuchi, and
M Matsushima
December 1999,
Chemical & pharmaceutical bulletin,
T Yamada, and
H Song, and
K Inaka, and
Y Shimada, and
M Kikuchi, and
M Matsushima
January 2005,
Acta biomaterialia,
