NMR Studies Of Peptides Research Articles

Local Conformation and Dynamics of Isoleucine in the Collagenase Cleavage Site Provide a Recognition Signal for Matrix Metalloproteinases*

The mechanism by which enzymes recognize the "uniform" collagen triple helix is not well understood. Matrix metalloproteinases (MMPs) cleave collagen after the Gly residue of the triplet sequence Gly∼[Ile/Leu]-[Ala/Leu] at a single, unique, position along the peptide chain. Sequence analysis of types I-III collagen has revealed a 5-triplet sequence pattern in which the natural cleavage triplets are always flanked by a specific distribution of imino acids. NMR and MMP kinetic studies of a series of homotrimer peptides that model type III collagen have been performed to correlate conformation and dynamics at, and near, the cleavage site to collagenolytic activity. A peptide that models the natural cleavage site is significantly more active than a peptide that models a potential but non-cleavable site just 2-triplets away and NMR studies show clearly that the Ile in the leading chain of the cleavage peptide is more exposed to solvent and less locally stable than the Ile in the middle and lagging chains. We propose that the unique local instability of Ile at the cleavage site in part arises from the placement of the conserved Pro at the P(3) subsite. NMR studies of peptides with Pro substitutions indicate that the local dynamics of the three chains are directly modulated by their proximity to Pro. Correlation of peptide activity to NMR data shows that a single locally unstable chain at the cleavage site, rather than two or three labile chains, is more favorable for cleavage by MMP-1 and may be the determining factor for collagen recognition.

Conformational and self-assembly studies of helix forming hexapeptides containing two α-amino isobutyric acids

Single crystal X-ray diffraction studies reveal that three hexapeptides with general formula Boc–Ile–Aib–Xx–Ile–Aib–Yy–OMe, where Xx and Yy are Leu in peptide I, Leu and Phe in peptide II, and Phe and Leu in peptide III, respectively, adopt equivalent conformations that can be described as mixed 3 10/α-helice with two 4→1 and two 5→1 intramolecular N–H⋯O C H-bonds. The peptides do not generate any helix-terminating Schellman motif despite having Aib at the penultimate position from C-terminus. In the crystalline state, the helices are packed in head-to-tail fashion through intermolecular hydrogen bonds to create supramolecular helical structures. The CD studies of the three hexapeptides in acetonitrile indicate that they are folded in well-developed 3 10-helical structures. NMR studies of peptide I in CDCl 3 also suggest the formation of a homogeneous 3 10-helical structure. The field emission scanning electron microscopic (FE-SEM) images of peptide II in the solid state reveal a non-twisted ribbon-like morphology, which is formed through lateral association of non-twisted filaments.

NMR studies of peptides and proteins associated with membranes.

AbstractHigh-resolution structural information about peptides and proteins associated with membranes now can be obtained from experimental studies involving both multidimensional solution and high resolution solid-state NMR spectroscopy. The relatively recent development and application of NMR spectroscopy for studies of membrane proteins takes advantage of the two well-characterized model membrane environments of detergent micelles and phospholipid bilayers (Fig. 1). Representation of model membranes. (A) Micelle, (B) bilayer. KeywordsDipolar CouplingPituitary Adenylate Cyclase Activate PolypeptideRotational ResonanceSodium Dodecyl Sulfate MicelleChemical Shift TensorThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

NMR studies of peptides derived from the putative binding regions of cartilage proteins. No evidence for binding to hyaluronan.

Previous work has implicated sequences within the tandem repeats of cartilage link protein in the interaction of link protein with hyaluronan. This conclusion was based on competitive inhibition experiments using synthetic peptides (Goetinck, P. F., Stirpe, N. S., Tsonis, P. A., and Carlone, D. (1987) J. Cell Biol. 105, 2403-2407). Further investigation of this system using high resolution nuclear magnetic resonance, circular dichroism, and competitive inhibition with other peptides indicates that the previously observed inhibition of link protein-hyaluronan binding was not caused by peptide-hyaluronan interactions. Instead, nonspecific aggregation of the peptides with link protein is proposed to account for all of the experimental data. Consequently, there is no direct experimental evidence to support the conclusion that these sequences in the tandem repeats of link protein are responsible for the link protein-hyaluronan interaction. If these peptides do represent the hyaluronan binding regions of link protein, these results imply a highly structure-dependent interaction between link protein and hyaluronan. Conformational analysis of the peptides using two-dimensional nuclear Overhauser spectroscopy indicates that the linear peptides do not adopt any stable secondary structure. However, several residues in the disulfide-looped peptides exhibit connectivities, suggesting a relatively long-lived extended chain conformation, consistent with predictions of secondary structure based on sequence analysis.