2RBI

STRUCTURE OF BINASE MUTANT HIS 101 ASN

Summary for 2RBI

• Entry DOI: 10.2210/pdb2rbi/pdb
• Descriptor: RIBONUCLEASE (2 entities in total)
• Functional Keywords: endoribonuclease, microbial extracellular ribonuclease, phosphodiester transferase, hydrolase
• Biological source: Bacillus intermedius
• Cellular location: Secreted: P00649
• Total number of polymer chains: 2
• Total formula weight: 24407.16

Authors

Offen, W.A.,Okorokov, A.L. (deposition date: 1996-11-12, release date: 1997-03-12, Last modification date: 2024-04-03)

Primary citation

Okorokov, A.L.,Panov, K.I.,Offen, W.A.,Mukhortov, V.G.,Antson, A.A.,Karpeisky, M.Y.a.,Wilkinson, A.J.,Dodson, G.G.
RNA cleavage without hydrolysis. Splitting the catalytic activities of binase with Asn101 and Thr101 mutations.
Protein Eng., 10:273-278, 1997

PubMed Abstract: Members of the microbial guanyl-specific ribonuclease family catalyse the endonucleolytic cleavage of single-stranded RNA in a two-step reaction involving transesterification to form a 2',3'-cyclic phosphate and its subsequent hydrolysis to yield the respective 3'-phosphate. The extracellular ribonuclease from Bacillus intermedius (binase, RNase Bi) shares a common mechanism for RNA hydrolysis with mammalian RNases. Two catalytic residues in the active site of binase, Glu72 and His101, are thought to be involved in general acid-general base catalysis of RNA cleavage. Using site-directed mutagenesis, binase mutants were produced containing amino acid substitutions H101N and H101T and their catalytic properties towards RNA, poly(I), poly(A), GpC and guanosine 2',3'-cyclic phosphate (cGMP) substrates were studied. The engineered mutant proteins are active in the transesterification step which produces the 2',3'-cyclic phosphate species but they have lost the ability to catalyse hydrolysis of the cyclic phosphate to give the 3' monophosphate product.

PubMed: 9153077
DOI: 10.1093/protein/10.3.273

Experimental method

X-RAY DIFFRACTION (2.2 Å)