Obtaining Overstable Methionine Aminopeptidase for the Removal of Methionine From Recombinant Proteins
https://doi.org/10.33647/2713-0428-19-3E-47-51
Abstract
Cleavage of the N-terminal initiating methionine (Met1) is a critical coand post-translational modification affecting 50–70% of cellular proteins. During the production of recombinant proteins in the heterologous system of E. coli expression, Met1 cleavage often fails to occur, which leads to heterogeneity of the preparations obtained, changes in their activity and stability. This problem can be solved by treating recombinant proteins in vitro with a specific enzyme, methionine aminopeptidase (MAP). Currently available MAPs exhibit limited specificities and reaction conditions. We cloned a MAP from a hyperthermophilic bacterium, developed a method for enzyme purification, and studied a number of physicochemical properties. The new MAP enzyme is resistant to elevated temperatures. The MAP maintains a stable native state in a pH range from 3 to 11 units. The novel MAP enzyme can be used to remove N-terminal Met1 from recombinant proteins in vitro over a wide pH range and at elevated temperatures.
Keywords
methionine aminopeptidase,
Thermus thermophilus,
heterologous expression,
N-terminal methionine excision
Supporting agencies: the research was supported by a grant from the Russian Science Foundation, No. 23-24-00563
About the Authors
Yu. S. Lapteva
Institute for Biological Instrumentation of the Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences
Russian Federation
Russian Federation
Yulia S. Lapteva - Cand. Sci. (Biol.).
142290, Moscow Region, Pushchino, Nauki Ave., 3
V. V. Bykov
Institute for Biological Instrumentation of the Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences
Russian Federation
Russian Federation
Vyacheslav V. Bykov
142290, Moscow Region, Pushchino, Nauki Ave., 3
M. V. Trunilina
Institute for Biological Instrumentation of the Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences
Russian Federation
Russian Federation
Maria V. Trunilina
142290, Moscow Region, Pushchino, Nauki Ave., 3
I. S. Boldaevsky
Institute for Biological Instrumentation of the Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences
Russian Federation
Russian Federation
Igor S. Boldaevsky
142290, Moscow Region, Pushchino, Nauki Ave., 3
T. A. Kudryashov
Institute for Biological Instrumentation of the Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences
Russian Federation
Russian Federation
Timofey A. Kudryashov
142290, Moscow Region, Pushchino, Nauki Ave., 3
А. A. Vologzhannikova
Institute for Biological Instrumentation of the Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences
Russian Federation
Russian Federation
Alisa A. Vologzhannikova - Cand. Sci. (Biol.).
142290, Moscow Region, Pushchino, Nauki Ave., 3
A. S. Sokolov
Institute for Biological Instrumentation of the Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences
Russian Federation
Russian Federation
Andrey S. Sokolov - Cand. Sci. (Biol.).
142290, Moscow Region, Pushchino, Nauki Ave., 3
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Review
For citations:
Lapteva Yu.S., Bykov V.V., Trunilina M.V., Boldaevsky I.S., Kudryashov T.A., Vologzhannikova А.A., Sokolov A.S. Obtaining Overstable Methionine Aminopeptidase for the Removal of Methionine From Recombinant Proteins. Journal Biomed. 2023;19(3E):47-51. (In Russ.) https://doi.org/10.33647/2713-0428-19-3E-47-51
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