1M07

RESIDUES INVOLVED IN THE CATALYSIS AND BASE SPECIFICITY OF CYTOTOXIC RIBONUCLEASE FROM BULLFROG (RANA CATESBEIANA)

Summary for 1M07

• Entry DOI: 10.2210/pdb1m07/pdb
• Related: 1KM8
• Descriptor: 5'-D(*AP*CP*GP*A)-3', Ribonuclease (3 entities in total)
• Functional Keywords: rc-rnase-d(acga), ribonuclease, bullfrog, cytotoxicity, hydrolase-dna complex, hydrolase/dna
• Biological source: Rana catesbeiana (bullfrog)
• Total number of polymer chains: 4
• Total formula weight: 27318.37

Authors

Leu, Y.-J.,Chern, S.-S.,Wang, S.-C.,Hsiao, Y.-Y.,Amiraslanov, I.,Liaw, Y.-C.,Liao, Y.-D. (deposition date: 2002-06-12, release date: 2003-01-21, Last modification date: 2024-11-13)

Primary citation

Leu, Y.-J.,Chern, S.-S.,Wang, S.-C.,Hsiao, Y.-Y.,Amiraslanov, I.,Liaw, Y.-C.,Liao, Y.-D.
Residues involved in the catalysis, base specificity, and cytotoxicity of ribonuclease from Rana catesbeiana based upon mutagenesis and X-ray crystallography
J.Biol.Chem., 278:7300-7309, 2003

PubMed Abstract: The Rana catesbeiana (bullfrog) ribonucleases, which belong to the RNase A superfamily, exert cytotoxicity toward tumor cells. RC-RNase, the most active among frog ribonucleases, has a unique base preference for pyrimidine-guanine rather than pyrimidine-adenine in RNase A. Residues of RC-RNase involved in base specificity and catalytic activity were determined by site-directed mutagenesis, k(cat)/K(m) analysis toward dinucleotides, and cleavage site analysis of RNA substrate. The results show that Pyr-1 (N-terminal pyroglutamate), Lys-9, and Asn-38 along with His-10, Lys-35, and His-103 are involved in catalytic activity, whereas Pyr-1, Thr-39, Thr-70, Lys-95, and Glu-97 are involved in base specificity. The cytotoxicity of RC-RNase is correlated, but not proportional to, its catalytic activity. The crystal structure of the RC-RNase.d(ACGA) complex was determined at 1.80 A resolution. Residues Lys-9, His-10, Lys-35, and His-103 interacted directly with catalytic phosphate at the P(1) site, and Lys-9 was stabilized by hydrogen bonds contributed by Pyr-1, Tyr-28, and Asn-38. Thr-70 acts as a hydrogen bond donor for cytosine through Thr-39 and determines B(1) base specificity. Interestingly, Pyr-1 along with Lys-95 and Glu-97 form four hydrogen bonds with guanine at B(2) site and determine B(2) base specificity.

PubMed: 12499382
DOI: 10.1074/jbc.M206701200

Experimental method

X-RAY DIFFRACTION (1.8 Å)