Proteomics 6, 973C986 [PubMed] [Google Scholar] 47

Proteomics 6, 973C986 [PubMed] [Google Scholar] 47. all three enzymes contributed to the folding of the ribbon isomer of -ImI. Here, we identify this alternative disulfide-linked species in the venom of isomerase (PPI), and immunoglobulin-binding protein (BiP). Polypeptides containing one or more disulfides in their native state are likely to become substrates for PDI, the enzyme that catalyzes the oxidation, isomerization, and reduction of disulfide bonds. PDI contains four thioredoxin-like domains, two of which have the catalytic Cisomerization of peptidyl-prolyl bonds can impede folding of proline-containing proteins. Peptide bonds to proline are synthesized in the conformation. Although the majority of these bonds remain in isomerization. The enzyme that catalyzes this otherwise slow reaction is PPI, a ubiquitous protein that is present in almost all cellular compartments (6). In mammalian cells, a number of ER-resident PPIs have been identified, including PPI B. Interestingly, in addition to accelerating folding rates via isomerization of peptide bonds to proline, PPI was also shown to improve the efficiency of PDI-mediated folding of ribonuclease T1 by providing partially folded protein chains with the correct proline isomers (7). Similarly, oxidative folding rates of maurotoxin, a disulfide-rich peptide from scorpion venom, were highest in the presence of PDI and FKBP-12, a PPI isoform located in the cytosol (8). Another enzyme known to cooperate with PDI in the folding of disulfide-containing proteins is BiP, a member of the heat shock protein 70 (Hsp70) family. BiP binds to unfolded and partially misfolded proteins upon their entry into the lumen of the ER and limits protein misfolding and aggregation. It is also a key enzyme in the retrograde transport of misfolded proteins from the ER to the proteasome (9). In recent years, a number of studies have addressed the potential cooperative folding of disulfide-containing proteins by PDI and BiP. Investigations into the folding Mouse monoclonal to CD21.transduction complex containing CD19, CD81and other molecules as regulator of complement activation of antibody chains revealed that both enzymes cooperatively act in the refolding of Fab fragments (10). It has been suggested that BiP binds the unfolded polypeptide chains and keeps them in a conformation in which the cysteine residues are accessible for PDI (10). Immunoprecipitation experiments further revealed that PDIA6, a member of the PDI family, forms a non-covalent complex with BiP and shows specificity toward BiP client proteins in human fibroblast cells (11). Subsequently, the interaction between another PDI family member, PDIA3, and BiP was shown to improve folding rates of ribonuclease B and -lactalbumin (12). Together, these findings indicate that members of the PDI family can recruit or are themselves recruited by other Lasofoxifene Tartrate ER-resident enzymes and chaperones, such as BiP and/or PPI B, to cooperatively act in the oxidative folding of at least some client substrates. Recently, an ER-resident Lasofoxifene Tartrate complex comprising PDI, PPI B, and BiP was identified in human hepatoma and mouse lymphoma cells (13), reflecting tightly regulated compartmentalization of protein folding in the ER. Although our understanding of enzyme-guided folding of proteins is constantly improving, comparatively Lasofoxifene Tartrate little is known about the folding and assembly of small, Lasofoxifene Tartrate disulfide-rich peptides. Disulfide-rich peptides are widely distributed in the plant and animal kingdoms, where they serve diverse functions. Examples include antimicrobial peptides, such as the defensins; various protease inhibitors; hormones, including insulin; and a wide range of neurotoxins found in animal venoms (Table 1). Predatory marine snails of the genus synthesize a great diversity of disulfide-rich peptides that often carry additional post-translational modifications, most commonly proline hydroxylations, -carboxylations, and C-terminal amidations (14). The diversity of peptides generated by cone snails is astonishing. Each of the 500C700 species of cone snail is estimated to synthesize hundreds of different peptides with distinct disulfide connectivities (15). With their vast structural diversity, conotoxins can therefore be regarded as model disulfide-rich peptides for understanding general mechanisms of peptide folding, including the formation of correct disulfide bonds. TABLE 1 Diversity of disulfide-rich peptides and enzymes implicated in their biosynthesis Open in a separate window * C-terminal amidation, O = hydroxyproline, prolines and hydroxyprolines are highlighted.