Cystine Peptides: The Intramolecular Antiparallel P-Sheet Conformation of a 20-Membered Cyclic Peptide Disulfide R. KISHORE, zyxwvu Molecular Biophysics Unit, Indian Institute zy of Science; S. RAGHOTHAMA, Sophisticated Instruments Facility, Indian Institute of Science; zyxw and P. BALARAM, zyx * Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560 012, India Synopsis zyx A 20-membered cyclic peptide disulfide Boc-C ys-Val- Aib- Ma-Leu-C ys-NHMe I I S S has been synthesized zyxwvu as a conformational model for disulfide loops of limited ring size. 'H-nmr studies at 270 MHz establish the presence of three intramolecular hydrogen bonds involving the Leu, Val, and methylamide NH groups in CDCl,. Evidence for peptide aggregation in CDCl, is also presented. A structural transition involving loosening of the hydrogen bond formed by the Val NH group is observed upon the measured addition of (CD,),SO to CDCl,. Hydrogen-bonding studies, together with unusually low field positions of the Cys(1) and Cys(6) C"H resonances and high JHNCmH values provide support for an intramolecular antiparallel p-sheet conformation, facilitated by a chain reversal at the Aib-Ala segment. Extensive nuclear Overhauser effect studies provide compelling evidence for the proposed conformation and also establish a type I' p-turn at the Aib-Ala residues, the site of the chain reversal. INTRODUCTION Small disulfide loops containing a limited number of intervening amino acids between the two linked cysteine residues -cys-(X),-c ys- I I S - S form an important structural element in proteins and polypeptide hormones. We have undertaken a systematic investigation of the conformational proper- ties of small disulfide loops varying in size from 11- (one spacer residue) to 20- (four spacer residues) membered rings. Earlier reports from this laboratory described the conformational analysis of 14-membered cyclic peptide disul- fide~,l-~ which assume importance in view of their occurrence at the active * To whom correspondence should be addressed. Biopolymers, Vol. 26,873-891 (1987) zyxwvu 0 1987 John Wiley & Sons, Inc. CCC oooS-3525/87/060873-19$04.00