Crystallization of a Complex between Ribonuclease T1 and 2'-Guanylic Acid

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Abstract
Ribonuclease TI was crystallized under various conditions. Form I crystals were produced by microdialysis against 53%(v/v) 2‐methyl‐2,4‐pentanediol in 0.01 M sodium acetate, 0.05% 2′‐guanylic acid (2′GMP) and 0.02% NaN3 (pH 6.2–7.2). These crystals are tetragonal, space group P41212 and contain two molecules per asymmetric unit; cell dimensions are a=b= 5.86 nm, c= 13.28 nm. Form IIa and form IIb crystals were obtained by microdialysis from a buffer of 0.01–0.05 M sodium acetate, 0.25–0.5%, 2′GMP, 0.02% NaN3 and 2–5 mM calcium acetate (pH 4.0–4.4) in the presence of 50–75% (v/v) 2‐methyl‐2,4‐pentanediol. These crystals are orthorhombic, space group P212121, and contain one molecule per asymmetric unit; cell dimensions are a= 4.66 nm, b= 5.02 nm, c= 4.04 nm (form I) and a= 4.44 nm, b= 5.00 nm, c= 4.03 nm (form II). Using high‐performance liquid chromatography, it could be shown for all crystal forms that 2′‐GMP is bound in the crystals. The molecular ratio between RNase T1 and 2′GMP was 0.9 for form II crystals and thus agreed with a 1:1 enzyme‐nucleotide complex. Heavy‐atom derivatives were produced with lead acetate for form IIa crystals and with uranyl acetate for from IIb crystals. Three‐dimensional X‐ray analysis of the RNase‐T1· 2′GMP complex is under way.