Tive series of domains that provide a supporting “stalk” that holds the functional domain(s) away in the cell surface (9). Isopeptide bonds, both Lys-Asn and Lys-Asp, now seem to be common within the Ig-like domains that make up the shafts, or stalks, of those structures, supplying tensile strength and stability along the length in the assembly (14). These bonds form spontaneously on protein folding; the hydrophobic environment lowers the pKa on the lysine residue, enabling its nucleophilic attack on SignificanceWe describe an unprecedented kind of intramolecular crosslink within a protein molecule, which we’ve located inside the repetitive domains of a cell surface adhesin in the Gram-positive organism Clostridium perfringens. From high-resolution crystal structures in the protein, coupled with MS, we show that these domains contain intramolecular ester bonds joining Thr and Gln side chains. These bonds are generated autocatalytically by a serine protease-like mechanism and give the lengthy, thin protein with tremendously enhanced mechanical strength and protection from proteolytic attack. The bonds supply an intriguing parallel together with the internal isopeptide bonds that stabilize Grampositive pili. Bioinformatics analysis suggests that these intramolecular ester bonds are widespread and frequent in cell surface adhesion proteins from Gram-positive bacteria.Author contributions: H.K., C.J.S., P.G.Y., and E.N.B. created study; H.K. performed investigation; H.K., C.J.S., P.G.Y., and E.N.B. analyzed information; and H.K., C.J.S., P.G.Y., and E.N.B. wrote the paper. The authors declare no conflict of interest. This article is really a PNAS Direct Submission. S.J.R. is often a guest editor invited by the Editorial Board.| protein stabilitystriking function of globular proteins is that regardless of the chemical diversity inherent inside the side chains of their constituent amino acids, chemical reactions involving these side chains are very uncommon. This could possibly be explained by evolutionary choice, which minimizes reactions that could prejudice proper protein folding.Salicylic acid (potassium) site Therefore, the only prevalent example of a covalent cross-link involving protein side chains would be the disulfide bond, which forms only in an proper redox atmosphere when two Cys residues are brought collectively by protein folding.104566-45-2 Chemscene Nonetheless, some surprising examples of unexpected crosslinks have already been brought to light by protein structure analysis or by the observation of uncommon spectroscopic or biophysical properties.PMID:23557924 Examples involve the Cys-Tyr bond in galactose oxidase (1), which offers a radical center; equivalent bonds in some catalases (2); the His-Tyr bond in cytochrome C oxidase (three); and also the exceptional chromophore of GFP (four). These, and other examples, arise via intramolecular reactions facilitated by particular regional environments. The recent discovery of isopeptide bonds joining Lys and Asn side chains within the proteins that make up pili around the Gram-positive bacterium Streptococcus pyogenes (5), too as on other Gram-positive pathogens (six), has highlighted a class of proteins in which intramolecular cross-links appear to become remarkably prevalent. It includes not simply Gram-positive pili but numerous other cell surface adhesins, generally known as microbial surface components recognizing adhesive matrix molecules (MSCRAMMs) (7). Examples from the latter include the collagen-binding A domain and repetitive B domains from the Staphylococcus aureus collagen-binding surface protein Cna (8, 9), the fibronectin-binding protein FbaB frompnas.
