I have been trying to confirm the Km of a substrate (which is 34 +/- 4 mM). This value was obtained in 50 mM MOPS, pH 6.3. I conducted my kinetics assay in a buffer of pH 7 and obtained a Km value in the 21.5. According to this paper, Fig. 2C, the normalized specific activity of the enzyme is about 70% at pH 6.3 and about 47% at pH 7. If I divide 34 mM at pH 6.3 by 0.7 (which should get me the optimum Km at the optimum pH of 5.5) and then multiply by 0.48, then I get 23 mM. However, the paper says Km is between 30 - 38 mM, so if I divide 30 and 38 separately by 0.7 and multiply by 0.47, I get 20 and 25.6 mM respectively. Because my value falls within this range, this then must mean that I have the right enzyme and the same result as the paper.
So my questions are:
When the paper says Km is "34 +/- 4 mM", can I assume that means the Km can be anywhere between 30 - 38 mM? I'm surprised to see how wide the range is. I assumed Km is usually just one value with a deviation of at most 0.1.
Do pH change Km values? I understand that pH changes the shape(s) of the enzyme and/or substrate. Therefore that must affect how much it wants to bind to the substrate. If the enzyme's desire to bind to a substrate decreases due to increase in pH, for example, that would mean more substrates are needed to surround the enzyme, thus increasing Km.
If pH does change Km, is this how I determine the Km value of a different pH value if the Km value at another pH is already known? I know that specific activity and Michaelis constant are different, but how much product can be converted per minute depends on how much the enzyme likes to bind to a substrate, which is represented by the Michaelis constant. Did my reasoning and calculation arrive at the right conclusion? If not, how is the calculation done?