Gelatin and starch as stabilizers of the coupled enzyme system of luminous bacteria NADH:FMN-oxidoreductase-luciferase

Тип публикации: статья из журнала

Год издания: 2014

Идентификатор DOI: 10.1007/s00216-014-7987-1

Ключевые слова: Bacterial luciferase, NADH:FMN-oxidoreductase, Bioluminescence, Stabilization of enzymes, Gelatin, Starch, bacterial protein, flavine mononucleotide reductase, luciferase, Aliivibrio fischeri, chemistry, enzymology, kinetics, luminescence, metabolism, Bacterial Proteins, FMN Reductase, Luciferases, Luminescent Measurements

Аннотация: We have studied the effects of a gel-like environment on the characteristics of enzyme preparations based on the coupled enzyme system of luminous bacteria, NADH:FMN-oxidoreductase-luciferase, to design a stable immobilizing reagent for bioluminescent analysis. Natural polymers, gelatin and starch, were used to create a viscous, structured microenvironment. The stability of the coupled enzyme system to such physical and chemical environmental factors as temperature, pH, and ionic strength in gelatin and starch-containing media was examined. It was shown that both gelatin and starch have a stabilizing effect on the enzymes of luminous bacteria under specific conditions. In particular, the enzymes' activity is increased twofold in the presence of 1 and 5 % of gelatin at 20 A degrees C and 25 A degrees C, respectively (temperatures lower than the gel point). Also, the acceptable pH range of the coupled enzyme system expands into the alkaline region and becomes 6.8-8.1. Stabilization at low ionic strength (0.01-0.06 mol L-1) is observed. At the same time, microenvironments based on either gelatin or starch do not change the enzymes' thermal inactivation rate constants in the temperature range from 25 to 43 A degrees C. Finally, gelatin and starch are suitable for development of a reagent for immobilization of enzymes which would be stable and resistant to physical and chemical environmental conditions.

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Издание

Журнал: ANALYTICAL AND BIOANALYTICAL CHEMISTRY

Выпуск журнала: Vol. 406, Is. 23

Номера страниц: 5743-5747

ISSN журнала: 16182642

Место издания: HEIDELBERG

Издатель: SPRINGER HEIDELBERG

Авторы

  • Bezrukikh Anna (Siberian Fed Univ, Inst Fundamental Biol & Biotechnol, Lab Bioluminescent Biotechnol, Krasnoyarsk 660041, Russia)
  • Esimbekova Elena (Siberian Fed Univ, Inst Fundamental Biol & Biotechnol, Lab Bioluminescent Biotechnol, Krasnoyarsk 660041, Russia; Inst Biophys SB RAS, Krasnoyarsk 660036, Russia)
  • Nemtseva Elena (Siberian Fed Univ, Inst Fundamental Biol & Biotechnol, Lab Bioluminescent Biotechnol, Krasnoyarsk 660041, Russia; Inst Biophys SB RAS, Krasnoyarsk 660036, Russia)
  • Kratasyuk Valentina (Siberian Fed Univ, Inst Fundamental Biol & Biotechnol, Lab Bioluminescent Biotechnol, Krasnoyarsk 660041, Russia; Inst Biophys SB RAS, Krasnoyarsk 660036, Russia)
  • Shimomura Osamu (Siberian Fed Univ, Inst Fundamental Biol & Biotechnol, Lab Bioluminescent Biotechnol, Krasnoyarsk 660041, Russia)

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