Isolation, Synthesis, and Antimicrobial Activities of Naturally Occurring θ-Defensin Isoforms from Baboon Leukocytes

Abstract

-Defensins are macrocyclic antimicrobial peptides that were previously isolated from leukocytes of a single species, the rhesus macaque. We now report the characterization of baboon -defensins (BTDs) expressed in bone marrow and peripheral blood leukocytes. Four cDNAs encoding -defensin precursors were characterized, allowing for the prediction of 10 theoretical -defensins (BTD-1 to BTD-10) produced by binary, headto-tail splicing of nonapeptides excised from paired precursors. Five of the predicted -defensins were purified from baboon leukocytes, and synthetic versions of each were prepared. Anti--defensin antibody localized the peptides in circulating neutrophils and monocytes and in immature and mature myeloid elements in bone marrow. Each of the BTDs possessed antimicrobial activity against bacterial and fungal test organisms in vitro. Peptide activities varied markedly despite a high degree of sequence conservation among the -defensins tested. Thus, baboons express numerous -defensins which appear to differentially contribute to host defense against diverse pathogens.Antimicrobial peptides (AMPs) are effectors of the innate immune system. AMPs are expressed in cells (epithelia, neutrophils, and macrophages) that come into contact with potentially invasive microorganisms (17). In mammals, the two major classes of AMPs are defensins and cathelicidins. Cathelicidins are characterized by a conserved cathelin prodomain which lies N terminal to highly variable mature peptides that are released by activating proteases (27). Defensins are small, cationic peptides that are composed of three structural subclasses, ␣-, ␤-, and -defensins, differentiated by the spacing and pairing of their six disulfide-bonded cysteines (7, 9, 18). -Defensins are further distinguished by their macrocyclic backbone and as such represent the only known cyclic protein motif expressed in animals (16).The biosynthesis of -defensins requires head-to-tail splicing of two 9-amino-acid sequences derived from -defensin precursors (16). -Defensins were first identified in neutrophils and monocytes of the rhesus monkey (21). Subsequently, Nguyen et al. (15) conducted a phylogenetic survey that revealed the existence of -defensin genes in other Old World monkeys and two apes (the siamang and orangutan), but none in New World monkeys or prosimians. Humans, chimpanzees, bonobos, and gorillas express -defensin pseudogenes in which the precursor mRNA contains a mutation producing a stop codon in the signal sequence, thus preventing translation of the -defensin precursor (15).Rhesus -defensin-1 (RTD-1) is produced from the heterodimeric splicing of two -defensin precursors, proRTD1a and proRTD1b. Homodimeric excision/ligation reactions involving proRTD1a and proRTD1b were revealed by the isolation of RTD-2 and RTD-3 (12, 23). RTD-1, -2, and -3 have potent microbicidal activities against bacteria and fungi (23) and have antiviral activities against human immunodeficiency virus type 1 (3, 24) and herpes simplex virus (26). A synthetic -defensin designed b...

Host defense peptides and their antimicrobial-immunomodulatory duality

“…␣-and ␤-Defensins are widely distributed across species, but ␥-defensins are only known to be expressed in granulocytes of the rhesus macaque and some other primates, including other Old World monkeys and orangutans (Crovella et al 2005;Selsted and Ouellette 2005). Other great apes including humans and NewWorld monkeys do not express ␥-defensins (Garcia et al 2008;Nguyen et al 2003;Selsted 2004;Tran et al 2008). ␥-Defensins are double-stranded small circular molecules, in contrast to ␣-and ␤-defensins, which are flat triple-stranded ␤ sheets (Boman 2003).…”

Interactions of α-, β-, and θ-Defensins with Influenza A Virus and Surfactant Protein D

“…-defensins, such as retrocyclins (RCs) or the RTD peptides found in rhesus macaque leukocytes, have a cyclic backbone and a unique, ladder-like pattern of disulfide bonds (5). Unlike ␣-and ␤-defensins, -defensin peptides are only expressed in nonhuman primates, such as the rhesus macaque and olive baboon (Papio anubis) (10,11). Humans express -defensin mRNA but lack the corresponding peptides because the human -defensin (DEFT) gene contains a stop codon in the signal sequence that aborts translation.…”

Biological Activities of Natural and Engineered Cyclotides, a Novel Molecular Scaffold for Peptide-Based Therapeutics

“…Daptomycin, is a 13-amino acid cyclic lipopeptide with a decanoyl side chain isolated from Streptomyces roseosporus that has recently been approved to treat infections against Gram-positive organisms, including multi-resistant strains [3]. In animals, the only known circular peptides are θ-defensins, which are expressed in blood leukocytes and bone marrow of Old World monkeys [4,5]. θ-Defensins are antimicrobial peptides with broad-spectrum activities against bacteria, fungi, and viruses [6][7][8].…”