O. Saether et al., ELUCIDATION OF THE PRIMARY AND 3-DIMENSIONAL STRUCTURE OF THE UTEROTONIC POLYPEPTIDE KALATA B1, Biochemistry, 34(13), 1995, pp. 4147-4158
The amino acid sequence and structure of a uterotonic polypeptide extr
acted from the African plant Oldenlandia affinis DC have been determin
ed. The peptide, kalata B1, consists of 29 amino acid residues and is
rich in cysteine (6), threonine (5), and glycine (5), Enzyme cleavage
studies show that the polypeptide backbone is cyclic. The three-dimens
ional solution structure has been determined using two-dimensional nuc
lear magnetic resonance (NMR) spectroscopy and distance-restrained sim
ulated annealing, Kalata B1 is composed mainly of beta-strands connect
ed by tight turns, forming regions of beta-sheet, except in the case o
f one section which forms a longer, less structured loop. The tertiary
fold, together with the disulfides that form a sulfur core, produces
a striking and unusual surface in which the majority of the hydrophobi
c residues form. a solvent-exposed patch. The hydrophobic side of kala
ta B1 is flanked by two diametrically opposed and opposite-charged res
idues. The structure calculations have been used to predict the previo
usly unknown disulfide bond connectivities using two approaches. In th
e first, a family of structures was calculated on the basis of NOE con
straints without the assumption of a specific disulfide connectivity.
The resultant structures were examined to determine whether the calcul
ated position of the sulfur atoms suggested that one set of disulfide
connectivities was more likely than the other, theoretically possible,
sets. In the second approach, a separate family of structures (50 per
set) was calculated for each of the 15 possible disulfide-bonded mole
cules. The resultant families of structures were compared to see wheth
er one was favored over the others, Both approaches led to the same gl
obal fold, and the most likely disulfide connectivity is predicted to
be 5-22, 13-27, and 17-29. In the calculated structure the cyclic pept
ide backbone is folded back onto itself and braced with disulfide pair
s across diagonally opposed beta-strands, This structure involves one
of the disulfide bonds (5-22) threading through the eight amino acid l
oop formed by the other two disulfide bonds and the peptide fragments
connecting them.