IFN-λ1 in CHO cells: its expression and biological activity.

BACKGROUND: Many studies have investigated the characteristics and biological activities of type III interferon (IFN), finding that it has similar features to type I IFN but also unique actions because it is recognized by a different receptor. RESULTS: A full-length recombinant human IFN-λ1 (rhIFN-λ1) cDNA was cloned into the pDF expression vector and stably expressed in Flp-In-CHO cells. After four purification steps (ammonium sulfate precipitation, SP Sepharose chromatography, Blue Sepharose 6 fast flow affinity chromatography and molecular sieve chromatography), the rhIFN-λ1 had a purity of about 90% and was found to have the predicted biological activities. The anti-viral activity of rhIFN-λ1 was determined as 106 IU/mg using the vesicular stomatitis virus (WISH-VSV) assay system. The anti-proliferation activity of rhIFN-λ1 was measured using the MTS method and the growth inhibition ratio was 57% higher than that for recombinant human IFN-α2b (rhIFN-α2b) when the rhIFN-λ1 concentration was 1000 IU/ml. rhIFN-λ1 had lower natural killer cell cytotoxicity than rhIFN-α2b. CONCLUSION: The Flp-In-CHO system is suitable for stably expressing rhIFN-λ1 that possesses the predicted anti-viral, anti-proliferation and natural killer cell cytotoxicity-promoting activities.

A triclinic polymorph with Z = 3 of N,N'-bis-(2-pyrid-yl)oxamide.

The asymmetric unit of the title compound, C(12)H(10)N(4)O(2), contains three half-mol-ecules. Each half-mol-ecule is completed by crystallographic inversion symmetry. The title compound, (I), is a polymorph of the structure, (II), reported by Hsu & Chen [Eur. J. Inorg. Chem. (2004), 1488-1493]. In the original report, the compound crystallized in the tetra-gonal space group P[Formula: see text]2(1)c (Z = 8), whereas the structure reported here is triclinic (P[Formula: see text], Z = 3). In both forms, each oxamide mol-ecule is almost planar (with maximum deviations are 0.266 and 0.166 Å) and the O atoms are trans oriented. The principal difference between the two forms lies in the different hydrogen-bonding patterns. In (I), two N-H⋯O and one N-H⋯N hydrogen bonds link the mol-ecules, forming a two-dimensional network, whereas in (II) there are no classical hydrogen bonds to O atoms and only weak C-H⋯O inter-actions are found along with rings of N-H⋯N bonds.

[Construction of human metapneumovirus DNA vaccine and study on its immune response in mice].

OBJECTIVE: To construct human metapneumovirus (hMPV) DNA vaccines and evaluate the cellular and humoral immune response in mice. METHODS: Fusion protein FdeltaTM (without transmembrane domain) gene and M gene of hMPV were amplified from cDNA by PCR, then DNA vaccines pcDNA3.1His-FdeltaTM and pcDNA3.1His-M were constructed to verify the expression of F and M protein by Western blotting and indirect immunofluorescent assay (IFA) respectively. Serum IgG and spleen cell CTL were detected with ELISA and ELISPOT assay after the BALB/c mice were immunized intramuscularly with the vaccines. RESULTS: The candidate DNA vaccines could express FdeltaTM and M protein as detected with Western blotting and IFA. The IgG antibody titers of mice was 1:44 when immunized with pcDNA3.1His-FdeltaTM, but could increase to 1:64 when co-immunized with pcDNA3.1His-M. ELISPOT assay demonstrated that IFN-gamma-secreting effector T cells reached 42 +/- 8.9 in co-immunization group, higher than single vaccine pcDNA3.1His-FdeltaTM group (32 +/- 7.4). CONCLUSION: DNA vaccine pcDNA3.1His-FdeltaTM could induce specific cellular and humoral immune responses, and the immune response could increase when co-immunization with pcDNA3.1His-M was carried out.

[Analysis of human metapneumovirus genotype in Hunan, China and its attachment protein G sequence character].

OBJECTIVE: To understand the genotypes of human metapneumovirus (hMPV) and the genetic character of hMPV attachment protein G sequence in Hunan, China. METHODS: 232 nasopharyngeal aspirates (NPA) samples from hospitalized children with acute respiratory infections were collected from Hunan, China in 2005. HMPV was detected. The full length of G glycoprotein genes were amplified and sequenced. Bioinformatics soft-wares were employed to analyze the sequences. RESULTS: 17/232 (7.3%) were showed hMPV positive. And co-infection rate with other viruses is 35%. The diagnoses of these hMPV positive cases are pneumonia, bronchiolitis and bronchopneumonia. Phylogenetic analysis for G genes from 13 hMPVs revealed the existence of four major subgroups: A1, A2, B1, B2 in Hunan, China in 2005. There are four types of sequence lengths of hMPV G glycoprotein, which are 711, 675, 660, 696nt. It is different in potential N-linked glycosylation sites and number of cysteine residues among these hMPVs of Hunan, China and Beijing, China. Also it is different from those in Japan and North America. CONCLUSION: The investigation of hMPV from Hunan, China in 2005 revealed the high speed of genetic variation and the marked character of geographic epidemic differences.

Diacetonitrile-1kappaN,3kappaN-bis{mu-trans-N-[3-(dimethylamino)propyl]-N'-(2-hydroxyethyl)oxamidato(2-)}-1:2kappa(5)N,N',O:O',N'';2:3kappa(5)O',N'':N,N',O-dithiocyanato-1kappaN,3kappaN-tricopper(II).

The title compound, [Cu(3)(C(9)H(17)N(3)O(3))(2)(NCS)(2)(CH(3)CN)(2)], contains two square-pyramidal Cu(II) units chelated by a transoid asymmetrical N-[3-(dimethylamino)propyl]-N'-(2-hydroxyethyl)oxamidate (dmapheoxd) dianion {H(2)dmapheoxd is N-[3-(dimethylamino)propyl]-N'-(2-hydroxyethyl)oxamide}, which coordinates to another Cu(II) ion in a square-planar environment lying on a crystallographic inversion center. Thus, the trans-oxamide ligand bridges two Cu(II) ions with different coordination numbers, and this is the first instance of such a zero-dimensional oxamide-bridged complex. The activated methyl group in the coordinated acetonitrile molecule is involved in a strong nonclassical C-H...O hydrogen bond, which contributes to a one-dimensional chain extending in the b direction. Considering the presence of weak bonding between the Cu atom and the uncoordinated hydroxyl O atoms, a two-dimensional structure is formed parallel to the ab plane.

Phospholipase A2-like activity of human bocavirus VP1 unique region.

Human bocavirus (HBoV) is a new parvovirus first discovered in 2005, which is associated with acute respiratory infection. Analysis of sequence homology has revealed that a putative phospholipase A2 (PLA2) motif exists in the VP1 unique region of HBoV. However, little is known about whether the VP1 unique region of HBoV has PLA2 enzymatic activity and how these critical residues contribute to its PLA2 activity. To address these issues, the VP1 unique region protein and four of its mutants, were expressed in Eschericha coli. The purified VP1 unique protein (VP1U) showed a typical Ca2+-dependent secreted PLA2-like (sPLA2) activity, which was inhibited by sPLA2-specific inhibitors in a time-dependent manner. Mutation of one of the amino acids (21Pro, 41His, 42Asp or 63Asp) in VP1U almost eliminated the sPLA2 activity of HBoV VP1U. These data indicate that VP1U of HBoV has sPLA2-like enzymatic activity, and these residues are crucial for its sPLA2-like activity. Potentially, VP1U may be a target for the development of anti-viral drugs for HBoV.