CONFORMATIONAL STUDIES OF AN UNDECAPEPTIDE REPRODUCING THE CONSENSUS SEQUENCE AROUND THE CLEAVAGE SITE OF THE RXVRG ENDOPROTEASE FROM XENOPUS-LAEVIS SKIN

Two synthetic fragments, corresponding to the 4-9 and 4-14 sequences o f a tetradecapeptide used as a model to test the RXVRG-endoprotease ac tivity from Xenopus laevis skin, have been studied by two-dimensional nmr spectroscopies, correlated spectroscopy, and nuclear Overhauser ef fect (NOE) spectroscopy. Both peptides wore the 5-9 consensus sequence found in several hormonal precursors. The nmr data for the 4-9 hexape ptide did not indicate any particular organization, either in water or in dimethylsulfoxide (DMSO), whereas, the 4-14 undecapeptide, a subst rate for the RXVRG endoprotease, showed, in DMSO solution, significant trends of structural organization involving the amino acids pertainin g to the consensus domain. From variations of integrated NOE peaks wit h temperature, the apparent interproton correlation times tau(c) were estimated and the maxima observed with Val7, the central residue in th e consensus sequence. A defined tertiary structure in that domain was also supported by medium- and long-range NOEs between Asp6 and Arg8, G lu4 and Gly9, and by the likely involvement of Arg8 and Gly9 NHs in in tramolecular hydrogen bonds. Most of these observations could be ratio nalized by an equilibrium between a 5-8 beta turn and a 9 --> 4 H-bond ed loop. The predominance of one rotamer for the C alpha-C beta bond w as established in four residues. Finally, the average Phi and Psi angl es were derived from two models taking, or not, into account variation s in the correlation times along the sequence. This allowed us to disc uss the artefacts generated by using an average correlation time throu gh the whole molecule. (C) 1994 John Wiley and Sons, Inc.