Structural and Functional Modeling of Human Lysozyme Reveals a Unique Nonapeptide, HL9, with Anti-HIV Activity

Abstract

We previously reported that lysozyme accounts for anti-HIV activity associated with the beta-core fraction of human chorionic gonadotropin [Lee-Huang, S., Huang, P. L., Sun, Y., Kung, H. F., Blithe, D. L. & Chen, H. C. (1999) Proc Natl Acad Sci U S A 96, 2678-81]. To define the structural and sequence requirements for anti-HIV activity, we carried out peptide fragmentation and activity mapping of human lysozyme. We identified two peptides that consist of 18 and 9 amino acids of human lysozyme (HL18 and HL9), corresponding to residues 98-115 and 107-115. HL18 and HL9 are potent inhibitors of HIV-1 infection and replication with EC(50)s of 50 to 55 nM, comparable to intact lysozyme. Scrambling the sequence or substitution of key arginine or tryptophan residues results in loss of antiviral activity. HL9, with the sequence RAWVAWRNR, is the smallest peptide we identified with full anti-HIV activity. It forms a pocket with its basic residues on the surface of the molecule. HL9 exists as an alpha-helix in native human lysozyme, in a region of the protein distinct from the muramidase catalytic site. Monte Carlo peptide folding energy minimizing simulation modeling and CD studies indicate that helical propensity does not correlate with antiviral activity. HL9 blocks HIV-1 viral entrance and replication, and modulates gene expression of HIV-infected cells, affecting pathways involved in survival, stress, TGFbeta, p53, NFkappaB, protein kinase C and hedgehog signaling.

Tonsil Epithelial Factors May Influence Oropharyngeal Human Immunodeficiency Virus Transmission

“…30 Gene expression analysis for innate immune molecules with anti-HIV potential such as SLPI, 5,36 ␤-defensins, 7 and thrombospondin 4 showed a trend for decreased expression in the tonsil epithelium. Of the molecules examined, only lysozyme, a broad-range antimicrobial factor with limited evidence of direct HIV targeting 37 was significantly up-regulated in the tonsil. SLPI, a protein with demonstrated anti-HIV activity in vivo and in vitro 46 was minimally expressed in the tonsil while abundant in the oral epithelia.…”

“…4 Interestingly, SLPI, defensins, and thrombospondin were expressed at a lower level in the tonsil epithelium (1.5-to 3-fold), although only SLPI was significantly different between tonsil and gingiva (P Ͻ 0.05). By comparison, lysozyme, a broad-range anti-viral factor with limited evidence of direct HIV activity, 37 was highly expressed in the tonsil (P Ͻ 0.05, Figure 5A). Immunohistochemical studies for SLPI revealed a striking difference of expression between tissues ( Figure 5B).…”

Cloning and functional expression of a human lysozyme gene (hly) from human leukocytes in Pichia pastoris

“…The addition of lysozyme to baby formulas was proposed (4), and lysozyme had also been shown to inhibit the growth of HIV-1 in vitro. Of note, the anti-HIV activity of lysozyme is independent of its muramidase activity (5). In addition, findings of previous studies have shown that it is effective in inactivating spores of Bacillus cereus in cheese by causing high hydrostatic pressure following the addition of lysozyme (6).…”

Lysozyme's lectin-like characteristics facilitates its immune defense function

“…In addition to the well-known muramidase activity of HL, increasing evidence suggests the existence of non-enzymatic and/or nonlytic modes of action against Gram-negative and Gram-positive bacteria (Lee-Huang et al 2005;Masschalck & Michiels, 2003). Furthermore, lysozyme has antitumor (Osserman et al 1973) and antiviral activities (Lee-Huang et al 2005), and it enhances the immune system (Siwicki et al 1998).…”

“…Furthermore, lysozyme has antitumor (Osserman et al 1973) and antiviral activities (Lee-Huang et al 2005), and it enhances the immune system (Siwicki et al 1998). The mechanisms of these activities remain unclear, and the dominant questions involve how HL recognizes pathogenic microbes.…”