Crystal structure of RNase T1(Y45W) complexed with 3'AMP and GflpA.
We have previously reported the crystallization of a mutant RNase T1(Y45W) with a synthetic modified trinucleotide ApGflpA [Hakoshima, T. et al. (1990) J. Biochem. 108, 695-698]. In the present report, we describe the crystal structure refined at 2.4 A resolution. During the refinement process, we found that the ApGflpA molecule was cleaved at the phosphodiester bond between the 5'-terminal adenosine and the subsequent 2'-fluoroguanosine. At the end of the refinement (R = 17.1%), it was supposed that the resulting molecules, i.e., 3'AMP and GflpA, were separately bound to the enzyme. In the complex structure, the binding-site of the enzyme was occupied by the guanine base of GflpA via a similar interaction to that of the enzyme complexed with 2'GMP, while the phosphate group of GflpA was not bound to the active site. The guanosine adopted the anti orientation on the glycosyl torsion angle with a C2'-endo-C3'-exo sugar pucker. This conformation resulted in the phosphate group protruding from the active site. The phosphate group of 3'AMP was bound to the active site of the enzyme and oriented itself toward the solvent region. This orientation was different from that of 2'AMP bound to the RNase T1(Y45W).
Gohda
K
K
Faculty of Pharmaceutical Sciences, Osaka University.
Itoh
T
T
Hiramatsu
Y
Y
Tomita
K
K
Nishikawa
S
S
Uesugi
S
S
Morioka
H
H
Ohtsuka
E
E
Ikehara
M
M
Hakoshima
T
T
D000241:Adenosine / Q000737:chemistry*; D000249:Adenosine Monophosphate / Q000737:chemistry*; D000595:Amino Acid Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000737:chemistry; D006147:Guanine / Q000378:metabolism; D006860:Hydrogen Bonding; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D006163:Ribonuclease T1 / Q000737:chemistry*
D000249:Adenosine Monophosphate; D006147:Guanine; C083670:2'-deoxy-2'-fluoroguanosine; D006163:Ribonuclease T1; D003849:Deoxyguanosine; D000241:Adenosine