As for most questions in science, the answer is unfortunately "it depends."
A substrate analog may bind just peachy at a pH in which the enzyme is
inactive, depending on what interactions are involved in stabilizing the
analog compared to the substrate transition state. Such a complex may
still be structurally informative.
Changing the ionization states of active site groups may or may not have
a significant effect on binding. One must consider not only
electrostatic interactions, but also hydrogen bonding donor-acceptor
interactions. Lowering pH, for example, may turn H-bonding acceptors
into obligate donors, or in the case of metalloenzymes, make metal-bound
water/hydroxide labile for exchange.
In any event, I'd be willing to bet that a substrate analog structure at
a non-optimal pH is more informative than no structure at the optimal pH! :)
Cheers,
_______________________________________
Roger S. Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346
tel: (315)-228-7245
ofc: (315)-228-7395
fax: (315)-228-7935
email: [log in to unmask]
On 10/30/2012 1:39 PM, Chittaranjan Das wrote:
> Peter,
>
> I think it would depend if the substrate analog have ionizable groups? If the analog does not have ionizable groups, it is hard to imagine how the the titration of ionizable groups on the protein would impair the binding.
>
> Chitta
>
>
>
> ----- Original Message -----
> From: "Peter Hsu" <[log in to unmask]>
> To: [log in to unmask]
> Sent: Tuesday, October 30, 2012 12:12:58 PM
> Subject: [ccp4bb] oof topic: pH effect on substrate analog
>
> Hi all,
>
> I'm working on a protein that I recently got crystals of. My functional studies show that the protein has optimal activity at lower pHs, while losing >90% activity at about pH8. I've been trying to soak/cocrystallize a substrate analog (small molecule) into my crystals (grown at ~pH8) with no real luck. I'm wondering, since I lack activity at this pH point, would it lead to no binding of a substrate analog?
>
> Thanks for any insights
>
> Peter
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