Selective and asymmetric action of trypsin on the dimeric forms of seminal RNase.
Dimeric seminal RNase (BS-RNase) is an equilibrium mixture of conformationally different quaternary structures, one characterized by the interchange between subunits of their N-terminal ends (the MXM form); the other with no interchange (the M=M form). Controlled tryptic digestion of each isolated quaternary form generates, as limit digest products, folded and enzymatically active molecules, very resistant to further tryptic degradation. Electrospray mass spectrometric analyses and N-terminal sequence determinations indicate that trypsin can discriminate between the conformationally different quaternary structures of seminal RNase, and exerts a differential and asymmetric action on the two dimeric forms, depending on the original quaternary conformation of each form. The two digestion products from the MXM and the M=M dimeric forms have different structures, which are reminiscent of the original quaternary conformation of the dimers: one with interchange, the other with no interchange, of the N-terminal ends. The surprising resistance of these tryptic products to further tryptic action is explained by the persistence in each digestion product of the original intersubunit interface.
De Lorenzo
C
C
Dipartimento di Chimica Organica e Biologica, Università di Napoli Federico II, Naples, Italy.
Dal Piaz
F
F
Piccoli
R
R
Di Maro
A
A
Pucci
P
P
D'Alessio
G
G
D000818:Animals; D002417:Cattle; D019281:Dimerization; D006868:Hydrolysis; D008297:Male; D013058:Mass Spectrometry / Q000379:methods; D008958:Models, Molecular; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D012260:Ribonucleases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D012661:Semen / Q000201:enzymology*; D014357:Trypsin / Q000737:chemistry / Q000378:metabolism*
D010446:Peptide Fragments; D012260:Ribonucleases; D014357:Trypsin