Journal
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
Volume 22, Issue 5, Pages -Publisher
MDPI
DOI: 10.3390/ijms22052768
Keywords
protein engineering; thermal stability; primary sequence; sequence profile comparison; adenylate kinase
Funding
- National Natural Science Foundation of China [11274076, 21773039]
- Shanghai Municipal Science and Technology Major Project [2018SHZDZX01]
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Protein engineering using rational design by bioinformatics can enhance thermostability of proteins without time-consuming experimental screenings. Utilizing a primary-sequence-based algorithm can help identify amino acid sites related to thermostability, leading to the construction of mutants with improved enzymatic activity.
Protein engineering is actively pursued in industrial and laboratory settings for high thermostability. Among the many protein engineering methods, rational design by bioinformatics provides theoretical guidance without time-consuming experimental screenings. However, most rational design methods either rely on protein tertiary structure information or have limited accuracies. We proposed a primary-sequence-based algorithm for increasing the heat resistance of a protein while maintaining its functions. Using adenylate kinase (ADK) family as a model system, this method identified a series of amino acid sites closely related to thermostability. Single- and double-point mutants constructed based on this method increase the thermal denaturation temperature of the mesophilic Escherichia coli (E. coli) ADK by 5.5 and 8.3 degrees C, respectively, while preserving most of the catalytic function at ambient temperatures. Additionally, the constructed mutants have improved enzymatic activity at higher temperature.
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