How do I inactivate Pepsin w/o using heat?

-BioDude1-

QUOTE (BioDude1 @ May 17 2007, 04:28 PM)

Can you adjust pH? Usually raising the pH to 7 will mostly inactivate it.

Jeff

-JVBishop-

The standard approach is to add 1 mM PMSF.

-phage434-

QUOTE (phage434 @ May 19 2007, 08:11 AM)

The standard approach is to add 1 mM PMSF.

Sorry, but PMSF is inhibitor of serine proteases mainly, isn't it? PMSF inibit trypsin, chymotrypsin, thrombin and papain.

We inhibit pepsin by heating at 80C and adjust pH till 7 or higher

-circlepoint-

QUOTE (BioDude1 @ May 17 2007, 09:28 PM)

How do I inactivate Pepsin w/o using heat?

fight the fire with fire: trypsination will do

-The Bearer-

Circlepoint is absolutely right. Sorry about that. It sounds as if Pepstatin A (Sigma P5318) would do the job at 1 uMolar. The inhibition is not permanent, however, and the material is stable for only 1 day at RT.

-phage434-

QUOTE (BioDude1 @ May 17 2007, 09:28 PM)

How do I inactivate Pepsin w/o using heat?

the question is, if "inactivation" should be permanent or not; working with pH shift to neutral and combination with inhibitors may not result in 100% permanent inactivation

-The Bearer-

QUOTE (The Bearer @ May 20 2007, 02:06 AM)

QUOTE (BioDude1 @ May 17 2007, 09:28 PM)

How do I inactivate Pepsin w/o using heat?

the question is, if "inactivation" should be permanent or not; working with pH shift to neutral and combination with inhibitors may not result in 100% permanent inactivation

I think that the goal of proteolytic digestion is to obtain protein fragments that futher analysed by MS or sequenator to identify targeted sequence or undergo futher purification with common chromatographic techniqes to obtain targeted peptides so question about inactivation means non permanent action.

-circlepoint-

How do I inactivate Pepsin w/o using heat? - (May/17/2007 )