Cys74 and Cys163 are necessary for IL-18 to elicit IFN-gamma production from peripheral blood lymphoid mononuclear cells.

There are four cysteines (Cys74, Cys104, Cys112 and Cys163) in mature human IL-18 (hIL-18). These cysteines are highly conserved in IL-18s of 11 species cloned so far, suggesting that one or more of the cysteines may be important for hIL-18 function. In this study, each cysteine residue was individually replaced with serine by site-directed mutagenesis.

[Interleukin-18 and its related cytokines in plasma of patients with idiopathic thrombocytopenic purpura].

To explore the role of immune regulating cytokines in pathogenesis of the idiopathic thrombocytopenic purpura (ITP) and its clinical significance, the levels of IL-18, TNF-alpha and Sc5b-9 in plasma of 32 ITP patients and 18 normal individuals were detected using ELISA methods. The results showed that IL-18, TNF-alpha and sC5b-9 levels in plasma of ITP patients were higher than that in normal individuals. The level of IL-18 was positively correlated with the levels of TNF-alpha and sC5b-9.

[Alteration of plasma interleukin-18 level in leukemia patients and its clinical significance].

To explore the alteration of plasma level of IL-18 in patients with leukemia before and after chemotherapy and its clinical significance, the plasma level of IL-18 was determined with ELISA method before and 2 weeks after chemotherapy in 37 leukemia patients, and 18 normal individuals. The results showed that the plasma IL-18 level (153.34 +/- 50.

Identification of important amino acid residues for human IL-18 function by mutant construction.

To study the structure-function relationship of IL-18, two IL-18 mutants, N- and C-terminal mutant (Delta NC) and IL-1 signature-like sequence mutant S(154)A/Y(156)F/E(157)P/C(163)T (S), were constructed by PCR. The wild type and mutant recombinant human interleukin-18 (rhIL-18) were expressed in E.coli, purified by Sephadex G-75 chromatography and renatured by stepwise dilution.

Cloning, High Level Expression and Purification of Rat betaB2-crystallin.

beta-crystallins are the largest group of lens structural proteins, which are necessary for both the high refractive index and the transparency of the eye lens, and have been implicated in various kinds of cataracts. To obtain abundant betaB2-crystallin for the study of the mechanism of their oligomerization, a bacterial expression system for betaB2-crystallin and a rapid method for its purification were developed. cDNA encoding rat betaB2-crystallin was cloned using RT-PCR.

Interleukin 18 High Level Expression in E.coli Purification and Renaturation of the Recombinant Protein.

Using the total RNA extracted from mitogen-stimulated human peripheral blood mononuclear cells (PBMC) as template, the cDNA of interleukin 18 was amplified by RT-PCR. The cDNA was subsequently cloned into the expression vector pJW2 and sequenced. The recombinant human IL-18 (rhIL-18) was expressed efficiently in inclusion bodies in E.

Asp(126), Asp(130) and Asp(134) are Necessary for Human IL-18 to Elicit IFN-gamma Production from PBMC.

To identify the amino acid residues which are critical to interleukin 18 (IL-18) function, three highly-conserved amino acids (Asp(126), Asp(130) and Asp(134)) were mutated to Asn, Lys and Lys. The wild type and mutant recombinant human interleukin-18 (rhIL-18) were expressed in E.coli, renatured by stepwise dilution and purified by Sephadex G-75 chromatography.

[Site-directed mutagenesis of the cysteines of human IL-18 and its effect on IL-18 activity].

To study the structure-function relationships of human IL-18(hIL-18), site-directed mutagenesis was used to generate four hIL-18 cysteine mutants, C74S, C104S, C112S and C163S. The cDNAs of the four cysteine mutants were inserted into prokaryotic expression vector pJW2 and expressed as inclusion bodies in E. coli.