The effect of Arg209 to Lys mutation in mouse thymidylate synthase

• Autorzy: Ciesla J. , Golos B. , Walajtys-Rode E. , Jagielska E. , Plucienniczak A. , Rode W.
• Języki publikacji: EN

Abstrakty

EN

Mouse thymidylate synthase R209K (a mutation corresponding to R218K in Lactobacillus casei), overexpressed in thymidylate synthase-deficient Escherichia coli strain, was poorly soluble and with only feeble enzyme activity. The mutated protein, incubated with FdUMP and N5,10-methylenetetrahydrofolate, did not form a complex stable under conditions of SDS/polyacrylamide gel electrophoresis. The reaction cata­lyzed by the R209K enzyme (studied in a crude extract), compared to that catalyzed by purified wild-type recombinant mouse thymidylate synthase, showed the Km value for dUMP 571-fold higher and Vmax value over 50-fold (assuming that the mutated en­zyme constituted 20% of total crude extract protein) lower. Thus the ratios kcat,R209K/kcat,'wild' and (kcat, R209K/Km, R209K dUMP)/ (kcat, 'wild'/Km, 'wild' dUMP) were 0.019 and 0.000032, respectively, documenting that mouse thymidylate synthase R209, similar to the corresponding L. casei R218, is essential for both dUMP binding and enzyme reaction.

Słowa kluczowe

EN

mouse; mice; thymidylate synthase; protein; 2'-deoxyuridylate; mutation; nucleotide

• Wydawca: -
• Czasopismo: Acta Biochimica Polonica
• Rocznik: 2002
• Tom: 49
• Numer: 3
• Opis fizyczny: p.651-658,fig.

Twórcy

autor

Ciesla J.

• Nencki Institute of Experimental Biology, L.Pasteura 3, 02-093 Warsaw, Poland

autor

Golos B.

autor

Walajtys-Rode E.

autor

Jagielska E.

autor

Plucienniczak A.

autor

Rode W.

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