Your cultures all finally got here. Thanks very much.
I'm glad to hear that "ONPG" is working well for you. I've been studying the coli K-12 enzyme in more detail,
with some very peculiar findings. The metal ions combine with the enzyme, and the major effect is not on the activity of the
enzyme, as determined at infinite substrate concentration, but on the Ks; in a formal way, the alkali metal ions "compete"
with the substrate, as well as with each other. This does not necessarily mean that they do absorb to the same sites on the
enzyme molecule, but they alter the affinity of the enzyme for the substrate in the same way that a competitive inhibitor
does (i.e. by increasing the slope of the 1/s term when 1/v is plotted against over 1/s). Lactose competes in accord with
the Lineweaver-Burk formulation; in M/50 Na phosphate buffer, the Ks is 1.3 x 10-4; KI (lactose) is about ten times higher.
Do you have some preparations with which the Nuerospora enzyme could be compared to the coli in these respects?
According to the grapevine, you've gotten several more lactose-negative mutants. Have you worked out a selective method?