It has long been appreciated that the photooxidation kinetics of amino acid (AA) residues in an intact protein differ from those of free AAs due to differences in the local steric microenvironment, such as its location in the three-dimensional structure. Yet there are only a few studies that have quantified the effect of protein structure on the photochemical reactivity of its residues. This is important for predicting phototransformation rates of AAs in aquatic environments where AAs in combined forms (e.g., oligopeptides and proteins) are more abundant than free AAs. In this work, the photochemical reactivity differences between free and combined AAs were assessed. Singlet oxygen (1O2) reaction kinetics of individual photooxidizable residues in the protein glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were examined. The results suggest that the 1O2 accessibility of residues in intact GAPDH has a profound effect on their photodegradation kinetics and for histidine residues can explain most of the variation in 1O2 reactivity. Additionally, 1O2-accessibile surface area values of residues calculated from protein crystal structure data are useful in predicting their reaction rates in GAPDH. This work illustrates a new approach to assess the differential photochemical reactivity of AA-based biomolecules in natural environments or engineered applications.
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