Model peptide studies of sequence repeats derived from the intracrystalline biomineralization protein, SM50. II. Pro,Asn-Rich tandem repeats

In the biomineralization process, a number of Pro-rich proteins participate in the formation of three-dimensional supramolecular structures. One such protein superfamily, the Pro,Gly-rich sea urchin intracrystalline spicule m atrix proteins, form protein-protein supramolecular assemblies that modify the microstructure of the inorganic mineral phase (calcite) within embryoni c sea urchin spicules and adult sea ui-chin spines. These proteins represen t ct useful model for understanding Pro sequence usage and the resulting ge neration of extended or "open" structures for protein-protein and/or protei n-crystal recognition. In the sea urchin spicule matrix protein, SM50 (Stro ngylocentrotus purpuratus), there exists an unusual 20-residue Pro,Asn-cont aining repeat, -PNNPNNPNPNNPNNPNNPNP-, which links the upstream 15-residue C-terminal domain and the downstream 211-residue beta -spiral repeat domain . To define the structural preferences of this 20-residue repent, rye creat ed a 20-residue N- and C-terminal "capped" peptidomimetic of this sequence. Using far-uv CD dichroism, CHalpha and alpha-N-15 conformational shifts, ( 3)J(NH-CH alpha) coupling constants, sequential d(NN(i, i + 1)) rotating fr ame nuclear Overhauser effect connectivities, d(alphaN(i, i + 1))/d(NN(i, i + 1)) intensity ratios, amide temperature shift coefficients, amide solven t exchange, and simulated annealing refinement protocols, we have determine d that this 20-residue repeat motif adopts an extended "twist" structure co nsisting of turn- and coil-like regions, These findings are consistent with previous studies, which have shown that Pro-rich tandem repeats adopt exte nded, flexible structures in solution. We hypothesize that this 20-residue repent may fulfill the role of a mineral-binding domain, a protein-protein docking domain, or as an internal "molecular spacer" for the SM50 protein d uring spicule biocomposite formation, (C) 2000 John Wiley & Sons, Inc.