Assignment and secondary-structure determination of monomeric bovine seminal ribonuclease employing computer-assisted evaluation of homonuclear three-dimensional 1H-NMR spectra.
Monomeric bovine seminal ribonuclease (mBS-RNase), the subunit of dimeric bovine seminal ribonuclease (BS-RNase), is an unusual monomer: for its structural stability, its catalytic activity, which is even higher than that of the parent dimeric enzyme, and for its role as an intermediate in the refolding of dimeric BS-RNase. Here we present the proton NMR assignment and secondary-structure determination of mBS-RNase, with a comparison of its structure to the structure of its parent protein, and to the structure of RNase A, a homologue with more than 80% identity in amino acid sequence. Proton NMR assignment was performed using a computer-assisted procedure, through a partially automated analysis of homonuclear three-dimensional spectra [Oschkinat, H., Holak, T. A. & Cieslar, C. (1991) Biopolymers 31, 699-712]. The secondary structures of mBS-RNase, of the A chain of dimeric BS-RNase, and of RNase A, are found to be similar. Significant differences are found instead, between mBS-RNase and RNase A in the more flexible stretches of the molecule, where a higher number of substitutions is present. Furthermore, a preliminary tertiary-structure model is reported, showing that the overall folding of mBS-RNase is closer to that of RNase A rather than that of (dimeric) BS-RNase.
D'Ursi
A
A
Dipartimento di Chimica, Università di Napoli Federico II, Italy.
Oschkinat
H
H
Cieslar
C
C
Picone
D
D
D'Alessio
G
G
Amodeo
P
P
Temussi
P A
PA
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D006860:Hydrogen Bonding; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012259:Ribonuclease, Pancreatic / Q000737:chemistry; D012260:Ribonucleases / Q000737:chemistry*; D012661:Semen / Q000201:enzymology; D016415:Sequence Alignment
D012260:Ribonucleases; D012259:Ribonuclease, Pancreatic