Результаты исследований: Научные публикации в периодических изданиях › статья › Рецензирование
Characterization of the Altai Maral Chymosin Gene, Production of a Chymosin Recombinant Analog in the Prokaryotic Expression System, and Analysis of Its Several Biochemical Properties. / Belenkaya, S. V.; Bondar, A. A.; Kurgina, T. A. и др.
в: Biochemistry (Moscow), Том 85, № 7, 01.07.2020, стр. 781-791.Результаты исследований: Научные публикации в периодических изданиях › статья › Рецензирование
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TY - JOUR
T1 - Characterization of the Altai Maral Chymosin Gene, Production of a Chymosin Recombinant Analog in the Prokaryotic Expression System, and Analysis of Its Several Biochemical Properties
AU - Belenkaya, S. V.
AU - Bondar, A. A.
AU - Kurgina, T. A.
AU - Elchaninov, V. V.
AU - Bakulina, A. Yu
AU - Rukhlova, E. A.
AU - Lavrik, O. I.
AU - Ilyichev, A. A.
AU - Shcherbakov, D. N.
N1 - Publisher Copyright: © 2020, Pleiades Publishing, Ltd. Copyright: Copyright 2020 Elsevier B.V., All rights reserved.
PY - 2020/7/1
Y1 - 2020/7/1
N2 - For the first time, the chymosin gene (CYM) of a maral was characterized. Its exon/intron organization was established using comparative analysis of the nucleotide sequence. The CYM mRNA sequence encoding a maral preprochymosin was reconstructed. Nucleotide sequence of the CYM maral mRNA allowed developing an expression vector to ensure production of a recombinant enzyme. Recombinant maral prochymosin was obtained in the expression system of Escherichiacoli [strain BL21 (DE3)]. Total milk-coagulation activity (MCA) of the recombinant maral chymosin was 2330 AU/ml. The recombinant maral prochymosin relative activity was 52955 AU/mg. The recombinant maral chymosin showed 100-81% MCA in the temperature range 30-50°C, thermal stability (TS) threshold was 50°C, and the enzyme was completely inactivated at 70°C. Preparations of the recombinant chymosin of a single-humped camel and recombinant bovine chymosin were used as reference samples. Michaelis–Menten constant (Km), turnover number (kcat), and catalytic efficiency (kcat/Km) of the recombinant maral chymosin, were 1.18 ± 0.1 µM, 2.68 ± 0.08 s−1 and 2.27± 0.10 µm M−1·s−1, respectively.
AB - For the first time, the chymosin gene (CYM) of a maral was characterized. Its exon/intron organization was established using comparative analysis of the nucleotide sequence. The CYM mRNA sequence encoding a maral preprochymosin was reconstructed. Nucleotide sequence of the CYM maral mRNA allowed developing an expression vector to ensure production of a recombinant enzyme. Recombinant maral prochymosin was obtained in the expression system of Escherichiacoli [strain BL21 (DE3)]. Total milk-coagulation activity (MCA) of the recombinant maral chymosin was 2330 AU/ml. The recombinant maral prochymosin relative activity was 52955 AU/mg. The recombinant maral chymosin showed 100-81% MCA in the temperature range 30-50°C, thermal stability (TS) threshold was 50°C, and the enzyme was completely inactivated at 70°C. Preparations of the recombinant chymosin of a single-humped camel and recombinant bovine chymosin were used as reference samples. Michaelis–Menten constant (Km), turnover number (kcat), and catalytic efficiency (kcat/Km) of the recombinant maral chymosin, were 1.18 ± 0.1 µM, 2.68 ± 0.08 s−1 and 2.27± 0.10 µm M−1·s−1, respectively.
KW - Michaelis–Menten kinetics parameters
KW - milk-clotting activity
KW - recombinant chymosin
KW - thermal stability
KW - CAMEL CHYMOSIN
KW - PASTORIS
KW - BOVINE CHYMOSIN
KW - Michaelis-Menten kinetics parameters
KW - ENZYME
KW - CLONING
KW - PURIFICATION
KW - PREPROCHYMOSIN
KW - ACTIVE CHYMOSIN
UR - http://www.scopus.com/inward/record.url?scp=85087873890&partnerID=8YFLogxK
U2 - 10.1134/S0006297920070068
DO - 10.1134/S0006297920070068
M3 - Article
C2 - 33040722
AN - SCOPUS:85087873890
VL - 85
SP - 781
EP - 791
JO - Biochemistry (Moscow)
JF - Biochemistry (Moscow)
SN - 0006-2979
IS - 7
ER -
ID: 24752889