ABSTRACT
Phylloquinone is a unique cofactor of photosystem I (PS I ) , made up of a redox-active naphthoquinone ring attached to a partially saturated C-20 phytyl side chain.At present, the research on the biosynthesis of phylloquinone in cyanobacteria is mainly focused on the formation of naphthoquinone ring, while there was a shortage of reports in the biosynthesis of phytyl side chain.In this study, a highly homologous protein S110875 was found in Synechocystis sp.PCC 6803 by homologous sequence alignment with VTE6, a kinase involved in phylloquinone biosynthesis by converting phytyl-phosphate into phytyl- diphosphate in Arabidopsis thaliana.The resulting S110875 mutant, called As/Z0875, accumulates none phylloquinone and tocopherol, as well as low amounts of chlorophyll content (P<0.05).The mutant had retarded growth in the absence of added glucose.Chlorophyll fluorescence, P700 absorbance changes, 77 K fluorescence emission spectra and Western blot analyses showed that in As/Z0875, PS I function was impaired and accumulation of the PS I complex was reduced remarkably (P<0.01), indicating that phvlloquinone deficiency affected PS I function, thus hindering the normal growth of cyanobacteria.Our results provide the evidence that the phytol phosphorylation pathway is essential for phylloquinone biosynthesis in cyanobacteria for the first time, and a basis for further investigate the protein synthesis, assembly and stability of PS I complex in cyanobacteria.
ABSTRACT
Eglin C is a small heat-stabilized protein from leeches. It belongs to the potato chymotrypsin inhibitor family, and can inhibit elastase, subtilisin, cathepsin, α-lytic protease and chymotrypsin etc. However, purification of protease by eglin C has not been studied. In this paper, the cDNA sequences encoding eglin C and its mutants were chemically synthesized and cloned into the prokaryotic expression vector pQE30. His
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the expression of transcription factors (TF) T-bet and GATA-3 mRNA in peripheral blood mononuclear cells and its correlation with immune status in esophageal cancer patients.</p><p><b>METHODS</b>Sixty patients were divided into two groups according to the clinical data: group A consisting of stage I and II, group B including stage III and IV. The gene expression of T-bet and GATA-3 in 60 esophageal cancer patients and 30 healthy controls was detected by reverse transcription-polymerase chain reaction (RT-PCR). The expression of IFN-gamma and IL-4 was measured by enzyme linked immunosorbent assay (ELISA).</p><p><b>RESULTS</b>Expression of T-bet mRNA in esophageal cancer patients (stage I and II: 0.27 +/- 0.05 ng/L, stage III and IV: 0.12 +/- 0.02 ng/L) was significantly lower than that in the healthy controls (1.35 +/- 0.14 ng/L), but the expression of GATA-3 mRNA in esophageal cancer patients (stage I and II: 0.45 +/- 0.06, stage III and IV: 0.55 +/- 0.03) was significantly higher than that in the healthy controls (0.09 +/- 0.10). The plasma level of Th1 cytokine IFN-gamma in the patients [stage I and II: (12.12 +/- 1.48) ng/L, stage III and IV: (8.44 +/- 0.90) ng/L] was significantly lower than that in the healthy controls, while the level of Th2 cytokine IL-4 in the patients [stage I and II: (18.64 +/- 0.77) ng/L, stage III and IV: (25.28 +/- 2.02) ng/L] was significantly higher than that in the healthy controls. However, neither in the expression of T-bet and GATA-3, nor in the plasma level of IFN-gamma and IL-4, showed a significant difference between group A and B.</p><p><b>CONCLUSION</b>In the peripheral blood of esophageal cancer patients, the expression of T-bet decreased, while GATA-3 increased, Th1/Th2 balance is broken, and the Th2 is dominant. T-bet and GATA-3 play a part role in the regulation of Th1/Th2 balance.</p>
Subject(s)
Adult , Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Enzyme-Linked Immunosorbent Assay , Esophageal Neoplasms , Allergy and Immunology , Metabolism , Pathology , GATA3 Transcription Factor , Genetics , Metabolism , Interferon-gamma , Blood , Interleukin-4 , Blood , Leukocytes, Mononuclear , Metabolism , Neoplasm Staging , RNA, Messenger , Metabolism , Reverse Transcriptase Polymerase Chain Reaction , T-Box Domain Proteins , Genetics , Metabolism , Th1 Cells , Metabolism , Th2 Cells , MetabolismABSTRACT
The effect of C-terminal region residues on the substrate specificity of a novel cyclic imide hydrolase (CIH), a recombinant cyclic imide hydrolase (CIH293), and its mutants deleted or substituted at C-terminus (CIH291, CIH290, KK292-293EE) was reported. The substrate specificity and kinetic parameters of the mutants were analyzed by both the spectrophotometric assay and high-performance liquid chromatography. Results show that the substrate specificity of mutants was not obviously changed, but slightly low for the affinity between the substrate and enzyme, compared with the wild-type enzyme, CIH293. In conclusion, the last three residues of CIH293 play an important role for the enzyme activity.