b-defensin-4

Antimicrobial Peptide / Chemoattractant for Monocytes

Three human b-defensins, with the conserved disulfide connectivity between Cys1-Cys5, Cys2-Cys4, and Cys3-Cys6 (Cys numbering from the amino-terminus), have been isolated thus far, yet 28 b-defensins were predicted in five gene clusters using a computational search approach [Proc. Natl. Acad. Sci. U.S.A., 99, 2129 (2002)]. These isolated peptides were actually human b-defensin-1 (hBD-1, Code 4337-s), hBD-2 (Code 4338-s ), and hBD-3 (Code 4382-s). Following these, the fourth b-defensin, hBD-4, was recently proposed based on the cDNA sequence analysis and the precursor of which is composed of 72 amino acid residues. Although natural hBD-4 has not yet been isolated, hBD-4 was tentatively designed as the peptide corresponding to the positions between 25 and 61 in the precursor sequence (hereafter the term "hBD-4" is used for this peptide and the following information is described in [FASEB J., 15, 1819 (2001)]). The sequence of hBD-4 has the b-defensin consensus core structure with 5 and 3 residue extensions from the amino- and carboxyl-terminal extreme cysteines in the core, respectively. Chemically synthesized hBD-4 was confirmed to share the conserved disulfide connectivity of the b-defensin family of peptides by the combination of enzymatic digestions and Edman degradation reaction.
b-defensin-4:
Glu-Leu-Asp-Arg-Ile-Cys-Gly-Tyr-Gly-Thr-
Ala-Arg-Cys-Arg-Lys-Lys-Cys-Arg-Ser-Gln-
Glu-Tyr-Arg-Ile-Gly-Arg-Cys-Pro-Asn-Thr-
Tyr-Ala-Cys-Cys-Leu-Arg-Lys
(Disulfide bonds between Cys⁶-Cys³³, Cys¹³-Cys²⁷,
and Cys¹⁷-Cys³⁴)

Using this chemically synthesized hBD-4, the following observations were reported: i) hBD-4 elicits the salt-sensitive antibacterial activities against both Gram-positive and Gram-negative bacteria in human respiratory epithelial cells; ii) the most active antibacterial activity is detected against Pseudomonas aeruginosa at 4.1 mg/ml; and iii) hBD-4 is a chemoattractant for human blood monocytes at 10 nM, but not for neutrophiles and eosinophiles. Interestingly, antibacterial activities in lung were inducible by the infection and subsequent activation of protein kinase C, thus differing from the activation mechanism from hBD-2 and hBD-3, which were induced in response to the stimulation by TNF-a, IL-1a, IL-6 or interferon a. hBD-4 mRNA was expressed abundantly in testis and stomach, and to a lesser extent but significantly in uterus, neutrophiles thyroid, lungs, and kidney.

hBD-4, which is regulated by specific stimulation that differs from those in hBD-2 and hBD-3, should be an essential component in clarifying the host defense mechanism in humans.