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, gelatin, luciferase, starch, Aliivibrio fischeri, chemistry, enzymology, kinetics, luminescence, metabolism, Aliivibrio fischeri, Bacterial Proteins, FMN Reductase, Gelatin, Kinetics, Luciferases, Luminescent Measurements, Starch

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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