Engineering disulfide bonds of the novel human β‐defensins hBD‐27 and hBD‐28: Differences in disulfide formation and biological activity among human β‐defensins

Abstract

Human β‐defensins comprise a large number of peptides that play a functional role in the innate and adaptive immune system. Recently, clusters of new β‐defensin genes with predominant expression in testicular tissue have been discovered on different chromosomes by bioinformatics. β‐Defensins share a common pattern of three disulfides that are essential for their biological effects. Here we report for the first time the chemical synthesis of the new fully disulfide‐bonded β‐defensins hBD‐27 and hBD‐28, and compare the results with synthetic procedures to obtain the known hBD‐2 and hBD‐3. While hBD‐27 was readily converted into a product with the desired disulfide pattern by oxidative folding, hBD‐28 required a selective protective group strategy to introduce the three disulfide bonds. The established synthetic processes were applied to the synthesis of hBD‐2, which, like hBD‐27, was accessible by oxidative folding, whereas hBD‐3 required a selective strategy comparable to hBD‐28. Experimental work demonstrated that trityl, acetamidomethyl, and t‐butyl are superior to other protection strategies. However, the suitable pairwise arrangement of the protective groups can be different, as shown here for hBD‐3 and hBD‐28. Determination of the minimum inhibitory concentration against different bacteria revealed that hBD‐27, in contrast to other β‐defensins tested, has virtually no antimicrobial activity. Compared to the other peptides tested, hBD‐27 showed almost no cytotoxic activity, measured by hemoglobin release of erythrocytes. This might be due to the low positive net charge, which is significantly higher for hBD‐2, hBD‐3, and hBD‐28. © 2004 Wiley Periodicals, Inc. Biopolymers (Pept Sci), 2005