0
$\begingroup$

I'm interested in any way to do time-resolved study of enzyme kinetics. I am studying some physical variables that may affect kinetics, but I want to study how quickly they take effect, and how long the effect lasts. The time scale of interest is milliseconds or less, so the typical spectrophotometric methods (which usually look at absorption change over fairly long times on the order of minutes) won't work.

Luciferase activity can be monitored by tracking the light emitted, and this has potentially very high time resolution as the time between reaction and light emission is on the order of microseconds. However luciferase is a bit of an odd model, as it is not part of normal metabolism or signalling in most organisms.

Is there a way to track enzyme kinetics of any other enzymes with a similarly fast time resolution?

EDIT Just discovered one name for what I'm looking for: "transient kinetics".

$\endgroup$
  • $\begingroup$ Is the change you want to observe visible in the UV/Vis range and you're asking for a way to observe this with very high resolution? $\endgroup$ – Mad Scientist Jun 22 '16 at 12:07
  • $\begingroup$ @MadScientist: I'm looking for good model systems, not any specific enzyme. The change of many enzymes is visible in UV/Vis, eg anything coupled to NAD/NADPH, unfortunately if doing a conventional progress curve type measurement the time resolution is more on the order of seconds or tens of seconds, not milliseconds. So it either has to be a somehow sped up way of doing progress curve, or some other way. $\endgroup$ – Alex I Jun 22 '16 at 20:41
  • 1
    $\begingroup$ Are you familiar with Stopped Flow? That sounds like what you're looking for. $\endgroup$ – Mad Scientist Jun 22 '16 at 22:12
1
$\begingroup$

Some ATPases can work with MANT-ATP or similar fluorescent ATP analogs that change their fluorescence properties upon protein binding as well as hydrolysis of their phosphate group. This has been used frequently (see here or here) to study enzyme kinetics on fast timescales.

$\endgroup$

Your Answer

By clicking “Post Your Answer”, you agree to our terms of service, privacy policy and cookie policy

Not the answer you're looking for? Browse other questions tagged or ask your own question.