Integrative structural biology: insights from photoactive yellow protein

The integration of computational approaches with multiple different experimental techniques offers an exciting avenue to address open questions in protein science. We report such an integrative structural biology approach using a bacterial photoreceptor (photoactive yellow protein) as an accessible model system. A coarse-grained model of PYP in its initial pG state and its light-activate pB intermediate was calibrated based on a range of experimental techniques and was used to obtain mechanistic and molecular insights into the structure and energetics of the pB intermediate. Engineered disulfide bonds were used to limit specific molecular motions in PYP. This approach revealed that the pB state is highly glassy, with multiple substates in which the N-terminal region of PYP adopts quite distinct conformations, which in turn greatly alter the lifetime of the pB intermediate. These results reconcile apparent contradictions in published literature regarding the structure of pB, demonstrate that the N-terminal region undergoes Functionally Important Motions (FIMs) during the PYP photocycle, and explains the classic observation that the N-terminal region of PYP modulates pB lifetime.