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1.
Front Microbiol ; 13: 1044226, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36439831

RESUMO

Glucose dehydrogenase (GDH) is a NAD(P)+ dependent oxidoreductase, which is useful in glucose determination kits, glucose biosensors, cofactor regeneration, and biofuel cells. However, the low efficiency of the catalysis hinders the use of GDH in industrial applications. In this study, an analysis of interactions between eight GDH mutants and NADP+ is powered by AlphaFold2 and Discovery Studio 3.0. The docking results showed that more hydrogen bonds formed between mutants, such as P45A and NADP+, which indicated that these mutants had the potential for high catalytic efficiency. Subsequently, we verified all the mutants by site-directed mutagenesis. It was notable that the enzyme activity of mutant P45A was 1829 U/mg, an improvement of 28-fold compared to wild-type GDH. We predicted the hydrophobicity of the protein-ligand complexes, which was confirmed by an 8-anilino-1-naphthalenesulphonic acid fluorescent probe. The following order of increasing hydrophobicity index was deduced: GDH < N46E < F155Y < P45A, which suggested that the enzyme activity of GDH is positively related to its pocket hydrophobicity. Furthermore, P45A still showed better catalytic ability in organic solvents, reaching 692 U/mg in 10% isopropanol, which was 19-fold that of the wild-type GDH. However, its substrate affinity was affected by organic solvents. This study provides a good theoretical foundation for further improving the catalytic efficiency of GDH.

2.
Artigo em Chinês | WPRIM (Pacífico Ocidental) | ID: wpr-664730

RESUMO

Objective To investigate the value of leukotriene B4(LTB4)and leukotriene E4(LTFA)in early diagnosis of renal involvement in children with Henoch Schonlein purpura(HSPN)and its influence on prognosis.Methods A total of 185 children with HSPN were enrolled in our hospital from January 2014 to October 2016.A total of 50 healthy children were selected as control group at the same period.The serum levels of LTB4 and LTFA in all subjects were detected,and their value in early diagnosis of HSPN and its influence on prognosis were analyzed.Results Compared with the control group,the serum levels of LTB4 and LT-FA in children with HSPN were significantly higher(P<0.05),the contents were 987.6 ng/L and 896.3 ng/L,respectively.The AUC of the combined detection of HSPN was 0.899 and the sensitivity was 0.810,which was better than that of LTB4 and LTB4 alone(P<0.05).The average follow-up was(28.4 ± 10.3)months,with grade A prognosis in 133 cases(71.9%),B grade in 47 ca-ses(25.4%),and C grade in 5 cases(2.7%).The rank sum test showed,the prognosis of children with LTB4<987.6 ng/L was better than that of children with LTB4≥987.6 ng/L,the prognosis of children with LTFA<896.3 ng/L was better than that of children with LTFA≥896.3 ng/L.Spearman analysis showed that the content of LTB4 and LTFA was negatively correlated with prognosis(Rs= -0.693 and -0.637,P<0.05).The higher contents of LTB4 and LTFA,the worse the prognosis.Conclusion Combined detection of LTB4 and LTFA has important clinical significance for early diagnosis and prognosis of HSPN.

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