X-RAY STRUCTURE DETERMINATION OF HUMAN GRO/MGSA PROTEIN. Hirofumi Ohishi*, Mitsuaki Sugawara#, Hironobu Nakayama#, and Ken-ichi Tomita*. Osaka University of Pharmaceutical Sciences, Matsubara, Osaka 580; #Faculty of Pharmaceutical Sciences,Osaka University, Suita, Osaka 565, Japan.

The human GRO (growth related protein)/ MGSA (melanoma growth stimulatory activity) protein with 73 amino acid (1) is a member of a super- family of chemotactic cytokineslike interleukin-8, platelet factor-4 (PF-4) monocyte chemoattractant factor and macrophage inflammatory protein-2. The human GRO gene was chemically synthesized and the secreted recombinant GRO protein expressed in E.coli was purified by chromatography, and used for crystallization by vapor diffusion. After trials with several different precipitants, the crystals grew as regular bipyramids with cubic cell dimensions: a=b=c=120.44 Å and space group I432, z=48 (dimeric GRO molecule per asymmetric unit). Intensity data up to 2 Å resolution were collected by RIGAKU R-AXIS imaging plate detector. The structure was solved by the molecular replacement method using the atomic coordinates for the dimer of PF-4(2) (from Protein Data Bank). and was then refined with 2.2 Å X-ray data, using CCP4, O and X- PLOR programs. The current overall basic structure found in GRO crystals is similar to the dimeric solution model by NMR spectroscopy(3), but the less restrained regions, the NH₂- and COOH-terminal regions are different from that of the PF-4 crystal structure and also of GRO NMR structure.

References:

1) A. Anisowicz et al., Proc. Natl. Acad. Sci. USA., 84, 718 (1987)

2) R. St. Charles et al., J. Biol. Chem., 264, 2092 (1989)

3) K-S. Kim et al., J. Biol. Chem., 269, 32909 (1994)