Abstract:  Single-molecule measurements are a valuable tool for revealing details of enzyme mechanisms by enabling observation of unsynchronized behavior.  However, this approach often requires immobilizing the enzyme on a substrate, a process which may alter enzyme behavior.  We apply a microfluidic trapping device to allow, for the first time, prolonged solution-phase measurement of single enzymes in solution.  Individual redox events are observed for single molecules of a blue nitrite reductase and are used to extract the microscopic kinetic parameters of the proposed catalytic cycle.  Changes in parameters as a function of substrate concentration are consistent with a random-sequential substrate binding mechanism.

Redox cycling and kinetic analysis of single molecules of solution-phase nitrite reductase

243rd National Meeting (2012)

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