Hello Rupinder,

Your question stands at the right foundation of enzymology indeed. Yes buffer is not necessary to get enzyme functioning but buffer is necessary to get reliable results as shown by history:

1) In 1902, the French physical chemist Victor Henri published the very first kinetic analysis of enzyme functioning working on so-called "invertase" (today glycoside hydrolases of the GH32 and GH100 families). He demonstrated that the curve describing variation of enzyme rates versus susbtrate concentration is hyperbolic and thus that an enzyme-substrate complex must occur during enzyme catalysis. All this work was carried out with non-buffered solutions! Despite this he was the first to express mathematically the variation of the rate of enzyme reaction against substrate concentration as such:

v = (k.S) / (1 + S/K1 + S/K2) 

All this without buffered solutions!

2) The German physician and biological chemist Leonor Michaelis and the Canadian physician Maud Menten in 1913 repeated Henri's work and got their wonderful results showing for the first time that the formation of the enzyme-substrate complex indeed is an equilibirum and expressing the equation as such:

v = (kcat.S) /  (Km + S)

and presenting a graphic easier way to measure both kcat and Km with the famous double-reciprocal plot 1/v = f (1/S).

All their work was carried out with buffered solutions and only this way enabled to get reliable results.

Best regards

Prof Philippe Urban

It can but pH is usually a very important variable for activity. Pure water is never experienced in nature so some ions are usually needed. Some typical buffers can actually inhibit though, I worked with almond b-glucosidase and Tris killed it dead.

Hello Rupinder,

Your question stands at the right foundation of enzymology indeed. Yes buffer is not necessary to get enzyme functioning but buffer is necessary to get reliable results as shown by history:

1) In 1902, the French physical chemist Victor Henri published the very first kinetic analysis of enzyme functioning working on so-called "invertase" (today glycoside hydrolases of the GH32 and GH100 families). He demonstrated that the curve describing variation of enzyme rates versus susbtrate concentration is hyperbolic and thus that an enzyme-substrate complex must occur during enzyme catalysis. All this work was carried out with non-buffered solutions! Despite this he was the first to express mathematically the variation of the rate of enzyme reaction against substrate concentration as such:

v = (k.S) / (1 + S/K1 + S/K2) 

All this without buffered solutions!

2) The German physician and biological chemist Leonor Michaelis and the Canadian physician Maud Menten in 1913 repeated Henri's work and got their wonderful results showing for the first time that the formation of the enzyme-substrate complex indeed is an equilibirum and expressing the equation as such:

v = (kcat.S) /  (Km + S)

and presenting a graphic easier way to measure both kcat and Km with the famous double-reciprocal plot 1/v = f (1/S).

All their work was carried out with buffered solutions and only this way enabled to get reliable results.

Best regards

Prof Philippe Urban

Thank you all...for your valuable suggestions.....

Dear friend. The enzymes have a very pH dependent catalytic activity profile. I.e., enzymes have a value of acidity in which its activity is maximum. You can check it. The blood has a pH that is very well regulated. If not, we would suffer alkalosis or acidosis. Both ends of great risk for sustaining life. Enzymes must be in a regulatory system in which, also, this activity is maximum. It could be in buffer phosphate or baking or tris. You can select and try among several regulators of pH in which increased activity have the enzyme that works.

A buffer is NOT to keep a solution neutral (at pH 7); its function is to minimize the change in pH when base or acid is added to the solution. Also note that there are many different buffers, and each one will stabilize the pH of a solution only within a specific pH range.

Therefore buffers are needed for enzymes for its stability as if pH changes during any processes then it also affect the enzymes a lot.

A nice question. All the answers so far are good - you should really follow up Prof Urban's comments. As far as my coments go - you should think that proteins retain their structures and activities in a cellular environment - with isolated proteins we have to TRY to mimic this - hence buffers and additives - like surfactants. Lipases, for an extreme example, appear to work best (adsorbed on beads) in solvents like hexane......but that is their natural environment - immobilised and in a hydrophobic medium. Compare like with like. A membrane enzyme and a cytoplasmic enzyme face different challenges.

Buffer provides a stable environment for the better action of an enzyme..

Prof Urban's and Robert 's comments are very good!

Actually we are trying to have a similar and stable environment for enzyme activity. Needless to say that different enzyme have different pH optima and they also demonstrate different pH stability. Therefore, we should carefully choose the best buffer and their pH for getting more accurate and reliable results. Otherwise, there will an awful lots of variations.

Good luck