The L1 major capsid protein of human papillomavirus (HPV) type 11, a 55-kDa polypeptide, forms particulate structures resembling native virus with an average particle diameter of 50 -60 nm when expressed in the yeast Saccharomyces cerevisiae. We show in this report that these virus-like particles (VLPs) interact with heparin and with cell-surface glycosaminoglycans (GAGs) resembling heparin on keratinocytes and Chinese hamster ovary cells. The binding of VLPs to heparin is shown to exhibit an affinity comparable to that of other identified heparin-binding proteins. Immobilized heparin chromatography and surface plasmon resonance were used to show that this interaction can be specifically inhibited by free heparin and dextran sulfate and that the effectiveness of the inhibitor is related to its molecular weight and charge density. Sequence comparison of nine human L1 types revealed a conserved region of the carboxyl terminus containing clustered basic amino acids that bear resemblance to proposed heparin-binding motifs in unrelated proteins. Specific enzymatic cleavage of this region eliminated binding to both immobilized heparin and human keratinocyte (HaCaT) cells. Removal of heparan sulfate GAGs on keratinocytes by treatment with heparinase or heparitinase resulted in an 80 -90% reduction of VLP binding, whereas treatment of cells with laminin, a substrate for ␣ 6 integrin receptors, provided minimal inhibition. Cells treated with chlorate or substituted ␤-D-xylosides, resulting in undersulfation or secretion of GAG chains, also showed a reduced affinity for VLPs. Similarly, binding of VLPs to a Chinese hamster ovary cell mutant deficient in GAG synthesis was shown to be only 10% that observed for wild type cells. This report establishes for the first time that the carboxyl-terminal portion of HPV L1 interacts with heparin, and that this region appears to be crucial for interaction with the cell surface.

The antiviral characteristics of monoclonal antibody IAM-41-2F5 (2F5) were determined in cell culture.

A notable feature of Ly-5, among immunogenetic systems that identify glycoproteins of the cell surface and define the surface phenotype of cells according to their lineage, is that the Ly-5 locus specifies a range of molecular isoforms that distinguish cells of different stages and branches of hematopoietic development. The composition of the Ly-5 locus is of much interest in regard to how these isoforms are constructed and differentially regulated according to cell lineage. We describe here a cDNA clone, pLy-5-68, that identifies Ly-5. The Ly-5 specificity of the pLy-5-68 clone was first indicated by a restriction fragment length polymorphism (RFLP), which in Southern blotting distinguishes genomic DNA of C57BL/6 (B6) mice (Ly-5a) from that of B6-Ly_5b congeneic mice whose genome is the same as B6 except for the segment of chromosome 1 that bears Ly_5b. For the following reasons it is unlikely that pLy-5-68 represents a gene linked to Ly-5 that was carried over with Ly_5b during serial backcrossing to make the B6&Ly_5b congeneic strain. In all mouse strains The Ly-S locus of the mouse (1, 2) specifies a set of different glycoprotein isoforms, each of which is expressed on the surface of cells of a particular lineage within the hematopoietic compartment of development (3). Of five NaDod-S04/PAGE bands derived from Ly-5 precipitated from cells of different hematopoietic lineages by Ly-5 antiserum or monoclonal antibody (3-5), two have been studied biochemically in some detail. These represent a 200-kDa isoform expressed by T cells and a 220-kDa isoform expressed by B cells. The size of the respective protein components of these two isoforms, which appear to be highly related to one another in two-dimensional peptide mapping, is 160 kDa and 190 kDa, and the Ly-5 locus, on chromosome 1 (unpublished data), is the site of the protein structural gene or genes for at least these two isoforms (6). The cDNA probe described here should help to elucidate the genetic basis of Ly-5 isoform diversity. MATERIALS AND METHODSPreparation of Sublibraries Based on cDNA Insert Size. A T-cell cDNA library, consisting of 3.8 x 105 independent transformants, was constructed by using the pcD vector of Okayama and Berg (7) and poly(A)+ mRNA of the Con-Astimulated (22 hr) inducer T-cell clone C1.Lyl-T1 (8). Plasmid DNA was prepared by the alkaline lysis procedure (9) followed by equilibrium sedimentation in CsCl. Twelve micrograms ofplasmid DNA was digested with 12 units of Sal I or Cla I endonuclease and electrophoresed in adjacent wells in a 1.3% agarose gel. DNA fragments whose sizes spanned the range 2-23 kilobases (kb) were electrophoresed in adjacent tracks. After staining with ethidium bromide, the gel was sliced into eight sections corresponding to cDNA insert sizes of 0-0.5, 0.5-1.2, 1.2-2, 2-3, 3-4, 4-5, 5-9, and 9-20 kb. DNA was extracted from each slice by the sodium iodide/glass powder method (10), recyclized with T4 DNA ligase, and used to transform Escherichia coli, strain MC1061. The individual transformed cultures...

No abstract

The Ly-5 system of the mouse is expressed exclusively by hematopoietic cells and comprises a series of glycoprotein isoforms that typify different hematopoietic cell lineages. The 200-kDa isoform of T cells and the 220-kDa isoform of B cells are known to differ in peptide composition. The complete 1152 amino acid sequence of the 200-kDa isoform protein deduced from cDNA sequence appears to comprise a leader sequence of some 30 residues, an external N-terminal domain of 370 residues, a probably single transmembrane domain of 22 residues, and an unusually large cytoplasmic domain of 730 residues. Both the external and cytoplasmic domains include regions of internal homology suggestive of evolution from a smaller ancestral gene. RNA transfer blotting has previously shown that B-cell mRNA for Ly-5 is larger than T-cell mRNA. S1 nuclease protection mapping with Ly-5 cDNA probes suggests that this difference can be ascribed to interpolation of an extra B-cell sequence located at the 5' end of B-cell mRNA, probably immediately following the leader sequence. From restriction mapping of overlapping Ly-5 genomic clones spanning 60 kilobases it is concluded that Ly-5 isoforms are generated by differential processing of transcripts of a single gene, rather than from a family of linked Ly-5 genes.