2
$\begingroup$

I need to know the concentration of pectinase enzyme (sigma aldrich) which has stated 5KU, 5U/mg protein (lowry) and lot result 20U/mg protein in the label of the enzyme bottle. This is all it has mentioned. How do I calculate the concentration from this information?

hey everyone, sorry for not been straight forward with the question. I'm asking for U/ml.I think I can find it if the label has mentioned the total volume of enzyme solution. Then I know how much mg in 5KU which is 5000/5=1000mg which I can easily get mg/ml or U/ml which is more appropriate. But the question is pointless if it hasn't mentioned the total volume. Thanks for the replies.

PS My supervisor asked me to prepare a standard graph of enzyme concentration (U/ml) against absorbance(O.D value) for DNS assay instead of a Galacturonic acid standard/glucose standard.(galacturonic acid is not available so he thinks it is better to you this graph than a standard glucose graph)

PS I checked the bottle again.On the bottom side of the bottle it has stated 10 ml.

Improve this question
$\endgroup$
4
  • $\begingroup$ What kind of concentration do you need? A mg/ml number? For enzymes this is relatively meaningless, since the activity is more important here (you can have a lot of dead protein in your preparation which is of no use for the enzyme reaction). So usually you get a U/ml (or whatever) concentration and you have a definition for the unit. $\endgroup$
    – Chris ♦
    Sep 27, 2015 at 15:49
  • $\begingroup$ Is it just me or anyone else find ironic that OP's name k.m and he/she is asking for enzyme concentration :P $\endgroup$
    – Dexter
    Sep 27, 2015 at 17:07
  • $\begingroup$ Welcome to BioSE! Please include a bit more information on how you have tried to solve the problem yourself. $\endgroup$
    – fileunderwater
    Sep 27, 2015 at 17:55
  • $\begingroup$ If you want to know the concentration in moles/litre then you need the molecular weight of the protein molecule--you did not provide this information. You would also need the volume of the solution in the bottle--you did not provide this information. moles = weight (in grams) / molecular weight. $\endgroup$
    – mdperry
    Sep 27, 2015 at 19:01

1 Answer 1

Reset to default
2
$\begingroup$

The enzyme unit U is not a measure of enzyme concentration but its activity, defined as the conversion of 1 micro mole ($\mu$mol) of substrate per minute (under a specified condition such as pH and T)

Absorbance (O.D. at a specific wavelength) of the enzyme is a measure of enzyme concentration, regardless of its activity. Depending on the unit of the extinction coefficient, Absorbance can be converted directly by Beer's Law to enzyme concentration, typically in mg/mL or in the standard mM.

5U/mg is the specific activity of pectinase, and the bottle contains a total of 5 kilo U (KU), meaning that there is a equivalent of 1g of active enzyme inside (often in powder). If you assume all pectinase molecules inside the bottle are active, when dissolved in 10mL water, you have a pectinase concentration of 100mg/mL. If the molecular weight of pectinase is 31kDa (31000 g/mol), then 100mg/mL is the same as 3.2mM in concentration.

By the way, Lowry protein assay measures enzyme concentration, not activity in U while DNS assay measures activity (of enzyme that produces reducing sugars), not enzyme concentration.

Improve this answer
$\endgroup$

You must log in to answer this question.

Not the answer you're looking for? Browse other questions tagged .