Genetic encoding of photocaged noncanonical amino acids provides a powerful tool to study protein functions through optical control but is not yet available for acidic amino acids.Herein,we report the first site-speci...Genetic encoding of photocaged noncanonical amino acids provides a powerful tool to study protein functions through optical control but is not yet available for acidic amino acids.Herein,we report the first site-specific genetic encoding of a photocaged glutamate,4-methoxy-7-nitroindolinyl caged glutamate(MNI-Glu),into recombinant proteins via an expanded genetic code through evolved EcLeuRS/tRNA pair.Using two enzymes as examples,we demonstrate that substituting the conserved-active-site glutamate of a secreted alkaline phosphatase and a protease HRV3C to MNI-Glu allows photoregulatory control of their enzymatic activities.Our approach is an important addition to the photocaged noncanonical amino-acid toolbox and provides a general method to photocontrol protein activity based on caging a critical glutamate.展开更多
基金This work was financially supported by National Natural Science Foundation of China(92253301,U22A20332,92156025,and 21922701)the National Key Research and Development Program of China(2022YFA0912403 and 2021YFA0909900)+1 种基金the Beijing Natural Science Foundation(JQ20034)to T.L and Peking UniversityInnovation Fund for Outstanding Doctoral Candidates of Peking University Health Science Center(71006Y2460)to X.L.
文摘Genetic encoding of photocaged noncanonical amino acids provides a powerful tool to study protein functions through optical control but is not yet available for acidic amino acids.Herein,we report the first site-specific genetic encoding of a photocaged glutamate,4-methoxy-7-nitroindolinyl caged glutamate(MNI-Glu),into recombinant proteins via an expanded genetic code through evolved EcLeuRS/tRNA pair.Using two enzymes as examples,we demonstrate that substituting the conserved-active-site glutamate of a secreted alkaline phosphatase and a protease HRV3C to MNI-Glu allows photoregulatory control of their enzymatic activities.Our approach is an important addition to the photocaged noncanonical amino-acid toolbox and provides a general method to photocontrol protein activity based on caging a critical glutamate.
