C. Di Bello

Identification of the paired basic convertases implicated in HIV gp160 processing based on in vitro assays and expression in CD4+ cell lines.

The human immunodeficiency virus HIV envelope glycoprotein gp160 is synthesized as an inactive precursor, which is processed into its fusiogenic form gp120/gp41 by host cell proteinases during its intracellular trafficking. Kexin/subtilisin-related endoproteases have been proposed to be enzyme candidates for this maturation process. In the present study, 1) we examined the ability of partially purified precursor convertases and their isoforms to cleave gp160 in vitro. The data demonstrate that all the convertases tested specifically cleave the HIV envelope glycoprotein into gp120 and gp41. 2) We demonstrated that a 19-amino acid model peptide spanning the gp120/gp41 junction is cleaved by all convertases at the same gp160 site as that recognized in HIV-infected cells. 3) In an effort to evaluate specific convertase inhibitors, we showed that the α1-antitrypsin variant, α1-PDX, inhibits equally well the ability of the tested convertases to cleave gp160 in vitro. 4) Three lymphocyte cell lines were screened by reverse transcription polymerase chain reaction in an effort to identify which are the convertases expressed in the most common HIV target, the CD4+ lymphocytes. The data demonstrate that furin, PC5/6, and the newly cloned PC7 are the main transcribed convertases, suggesting that these proteinases are the major gp160-converting enzymes in T4 lymphocytes.

Heparin enhances the furin cleavage of HIV-1 gp160 peptides

Infectious HIV‐1 requires gp160 cleavage by furin at the REKR511↓ motif (site1) into the gp120/gp41 complex, whereas the KAKR503 (site2) sequence remains uncleaved. We synthesized 41mer and 51mer peptides, comprising site1 and site2, to study their conformation and in vitro furin processing. We found that, while the previously reported 19mer and 13mer analogues represent excellent in vitro furin substrates, the present extended sequences require heparin for optimal processing. Our data support the hypothesis of a direct binding of heparin with site1 and site2, allowing selective exposure/accessibility of the REKR sequence, which is only then optimally cleaved by furin.

CCR5 N-terminus peptides enhance X4 HIV-1 infection by CXCR4 up-regulation

The HIV-1 envelope glycoprotein gp120 interacts consecutively with CD4 and CCR5 to mediate the entry of R5-HIV-1 strains into target cells. The N-terminus of CCR5, which contains several sulfated tyrosines, plays a critical role in gp120-CCR5 binding and, consequently, in viral entry. Here, we demonstrate that a tyrosine sulfated peptide, reproducing the entire N-terminal extracellular region of CCR5, its unsulfated analogue, and a point-mutated peptide are unable to inhibit R5-HIV-1 mediated infection, competing with the entire CCR5 in the formation of gp120-CD4-CCR5 complex. Surprisingly, these peptides show the capability of enhancing HIV-1 infection caused by X4 strains through the up-regulation of both CD4 and CXCR4 receptors.

Chromatographic behaviour of ureas, thioureas, biuret derivatives and related compounds

Abstract The separation conditions of some non-aromatic heterocyclic and related compounds are given. Some chromatic reactions used to localize the products are described and briefly discussed.

Anti-HIV-1 activity of CD4 synthetic oligopeptides representative of the putative gp120 binding site

Two CD4 oligopeptides, corresponding to residues (37–53) and (37–55) of the V1 domain of CD4, which recent structural studies propose as the most likely binding site of HIV-1 gp120, have been chemically synthesized by solid-phase techniques, modified by the addition of two side-chain protected cysteines at both termini and purified by HPLC. Their ability to inhibit the infectivity of human immunodeficiency virus type 1 (HIV-1) (HTLV-IIIB, RF and GB8 strains) in different cell lines was monitored by the production of progeny virus, p24 and reverse transcriptase activity in the culture supernatants and by electron microscopy. The results indicated that the peptides inhibited HIV-1 infectivity in a dose-dependent fashion without any detectable cytotoxicity.

Taxonomic position of some rana species in a dendrogram from their globin composition

Abstract 1. 1. The aminoacidic composition of the globin of the genus Rana has been analyzed. 2. 2. The resulting data and those from five other species of Ranidae , from one species of Pipidae and one of Bufonidae having been compared in pairs according to the formula of Harris and Teller gave origin to a “dissimilarity matrix”. 3. 3. In a “Phylo” computer program, consistent with Moores model, the data of the input matrix have been used to find the mutual taxonomic relations of the 11 anuran species examined.

Fiber-entrapped dipeptidyl aminopeptidase

The enzyme dipeptidyl aminopeptidase has been immobilized on cellulose triacetate fibers. The Michaelis-Menten parameters of the entrapped enzyme have been determined and compared to those of the literature for the native enzyme. The entrapped enzyme retains 13% of its activity toward synthetic substrate. The influence of mixing effects on the catalytic properties of the enzyme has been substantiated. The activity appears to be controlled by external mass-transfer.

β-Carbonylamides as chelating agents. N.M.R. determination of the equilibria formation with lanthanide metal ions

Abstract Complex formation of N-acetoacetylisopropylamide with europium(III) in methanol has been studied by N.M.R. spectroscopy. The stability constant indicates a fairly good binding power for such a ligand, and seems promising in view of the utilization of β-chetoamides as chelating agents for the study of the relationships between structure and function in biological systems.