Synthesis and comparative properties of two amide‐generating resin linkers for use in solid phase peptide synthesis
Well‐characterized resins of high purity are critical for effective solid phase peptide synthesis (SPPS). The quality of commercial (4‐methyl)benzhydrylamine‐resin (MBHA‐resin), used for the synthesis of peptide amides, is not consistent and residual ketone functionalities are frequently present. Such ketone or aldehyde impurities lead to the formation of acylation‐resistant deletion peptides in SPPS. To avoid these undesirable side reactions, we have optimized the preparation of two amide‐generating linkers, which, in combination with aminomethyl‐resin prepared directly from polystyrene resin, serve as alternatives to MBHA‐resin for peptide amide synthesis. Then we have explored their comparative properties in SPPS. Use of sonication in reductive amination facilitated the synthesis of both benzhydrylamine (BHA) and MBHA linkers. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd.
Total chemical synthesis of the enzyme sortase A(ΔN59) with full catalytic activity.
The enzyme sortase A is a ligase which catalyzes transpeptidation reactions.1, 2 Surface proteins, including virulence factors, that have a C terminal recognition sequence are attached to Gly5 on the peptidoglycan of bacterial cell walls by sortase A.1 The enzyme is an important anti-virulence and anti-infective drug target for resistant strains of Gram-positive bacteria.2 In addition, because sortase A enables the splicing of polypeptide chains, the transpeptidation reaction catalyzed by sortase A is a potentially valuable tool for protein science.3 Here we describe the total chemical synthesis of enzymatically active sortase A. The target 148 residue polypeptide chain of sortase AΔN59 was synthesized by the convergent chemical ligation of four unprotected synthetic peptide segments. The folded protein molecule was isolated by size-exclusion chromatography and had full enzymatic activity in a transpeptidation assay. Total synthesis of sortase A will enable more sophisticated engineering of this important enzyme molecule.
Use of a specific antiserum for renin detection in human kidney.
The use of anti-human renin antibodies made possible the intrarenal localization of renin in human kidney by immunofluorescence. In normal kidney, only some juxtaglomerular apparatus (JGA) were fluorescent. In these JGA, granular or diffuse fluorescence was only seen in afferent arterioles and was not present in all cells. In the ischemic areas of partially infarcted kidney, fluorescence was seen in all JGA and in interlobular arteries. In these arteries the most eccentric cells were often the most positive. In the nonischemic areas of the same kidneys, fluorescence was not seen in JGA, but was observed in proximal tubular cells, suggesting the reabsorption of filtered renin at this site.