Identification of the gene encoding vaccinia virus immunodominant protein p35.

The major envelope protein, p35, of vaccinia virus (VV; strain LIVP) was purified by extraction from virions with the non-ionic detergent Nonidet P-40. The protein was cleaved with CNBr. Four homogeneous peptides were isolated and their N-terminal amino-acid (aa) sequences determined.

Inhibitory effect of calf fetuin on the cytotoxic activity of LAK cell-derived factors and tumor necrosis factor.

A protein-inhibiting LAK cell-mediated cytotoxicity was isolated from a LAK cell-conditioned medium. The N-terminal amino acid sequence of this protein appeared to be identical to fetal calf fetuin. Pure isolated fetuin, as well as commercially available preparations of this protein, were shown to inhibit cytotoxic activity of both cytotoxic proteins released by LAK cells and TNF.

Contribution of membrane-associated cytotoxic proteins to cytolysis of L929 and K562 target cells mediated by LAK cells with different phenotypes.

We have demonstrated that the relative contribution of secreted and membrane-associated proteins to the cytotoxicity of LAK cells depended on LAK cell phenotype: the cytotoxicity of CD16+ CD8+ CD3- LAK cells was associated mainly with membrane-bound proteins, and the activity of CD3+ CD8+ CD16- LAK cells was due mainly to secreted soluble proteins. The cytotoxic activity of membrane fractions of LAK cells against cell line L929 was determined by 40-, 32- and 21-kDa proteins for LAK cells bearing NK-specific markers and due to proteins of 34 and 21 kDa in the case of 'CTL-like' LAK-cells.

The role of apoptotic and necrotic processes in cytolysis mediated by LAK cells with different phenotypes.

The role of necrotic and apoptotic pathways in cytolysis mediated by LAK cells was studied. The contribution of necrotic and apoptotic processes to cytolysis depends both on the LAK cells' phenotype and the type of target cells. CD16+/CD8+/CD3- LAK cells induced necrosis of K562 and L929 target cells.

Time-dependent changes of LAK cell phenotypes correlate with the secretion of different cytotoxic proteins.

Human lymphokine-activated killer (LAK) cells were generated from peripheral blood lymphocytes (PBL) of normal volunteers by interleukin-2 (IL-2) stimulation for 1-8 days. During the first 3 days the surface marker CD16 characteristic for natural killer (NK) cells was expressed and later the CD3 marker characteristic for cytotoxic T cells became predominant. The conditioned media of LAK cells collected after interaction of LAK cells with K562 target cells was chromatographically separated into two cytotoxic fractions: F1 and F2.

Studies on the N-terminal sequences of lectins isolated from the seeds of Butea frondosa.

Two lectin fractions (FI and FII) were obtained from seeds of Butea frondosa by affinity chromatography on a sorbent of macroporous glass coupled to the disaccharide alpha-D-GalNAc-(1----3)-beta-D-Gal. Both of these fractions, although different in their sugar specificity, were found on SDS-PAGE to consist of two polypeptide chains of 33 kDa and 35 kDa. In the native state the subunits associated to form a 250 kDa complex, possibly comprising four molecules of the 33 kDa polypeptide and four molecules of the 35 kDa polypeptide.

Amino acid sequence determination of vaccinia virus immunodominant protein p35 and identification of the gene.

A major immunodominant envelope protein of vaccinia virus (protein p35) was purified by extraction from virions with the nonionic detergent Nonidet P-40. The protein was cleaved with cyanogen bromide. Four homogeneous peptides were isolated and their N-terminal amino acid sequences determined.

Effect of antibodies to granular perforin and calmodulin on the activity of natural killer cytotoxic factor.

It was previously established that the cell death induced by natural killer cytotoxic factor (NKCF) is a complex calcium-dependent two-stage process. In the initial stage, during the first 30 min of incubation of NKCF with target cells, pore formation on the surface of the target is observed, together with temporary membrane damage. The second stage cytolytic activity occurs after a 3 h latent period, reaches maximum at 24 h, and leads to cell death.

Mesoderm-inducing factor from bovine amniotic fluid: purification and N-terminal amino acid sequence determination.

Bovine amniotic fluid contains a protein capable of inducing mesoderm formation from animal cap cells isolated from Xenopus laevis embryos at the late blastula stage. The procedure for purification of this protein is described and includes the following steps: CM-cellulose cation-exchange chromatography, chromatography on hydroxylapatite, reverse-phase hplc, size exclusion hplc, and a second reverse-phase hplc. A partially purified preparation of a mesoderm-inducing factor with high mesoderm-inducing activity was thus obtained.

The primary structure of Escherichia coli RNA polymerase. Nucleotide sequence of the rpoB gene and amino-acid sequence of the beta-subunit.

The combined structural study of proteins and of their corresponding genes utilizing the methods of both protein and nucleotide chemistry greatly accelerates and considerably simplifies both the nucleotide and protein structure determination and, in particular, enhances the reliability of the analysis. This approach has been successfully applied in the primary structure determination of the beta and beta' subunits of Escherichia coli DNA-dependent RNA polymerase and of their structural genes, yielding a continuous nucleotide sequence (4714 base pairs) that embraces the entire rpoB gene, the initial part of the rpoC gene and the intercistronic region, together with the total amino acid sequence of the beta subunit, comprising 1342 residues, and the N-terminal sequence of the beta' subunit (176 residues).