Figure 2
From: Regulation of homocysteine metabolism by Mycobacterium tuberculosis S-adenosylhomocysteine hydrolase
Role of His363 in activity and effect of SahH on homocysteine concentration in M. smegmatis.
(A) Role of His363 in SAH-hydrolytic activity.Activities of 1.74 μM each of holo SahH-WT and holo SahH-H363A were compared. Graph shows increase in OD at 415 nm with time when SahH-WT was used. No increase in OD at 415 nm was observed with SahH-H363A mutant. Error bars represent standard deviation (SD). (B) Role of His363 in SAH-synthetic activity. Activities of 4 μM each of holo SahH-WT and holo SahH-H363A were compared. Autoradiograph of TLC shows SAH synthesis in reactions containing SahH-WT while reduced product formation was seen in reactions containing SahH-H363A. Adenosine (Ado) and SAH spots are indicated by arrows. (C) Histogram representing intracellular homocysteine concentration in M. smegmatis MC2 4517 with and without the over-expression of Mtb-SahH. Homocysteine concentration was analyzed using UPLC and plotted as μmoles/1015 CFU. Homocysteine levels increased to about six times in cells over-expressing SahH (see also Table 1). Error bars represent standard deviation of three independent experiments and * p < 0.05 as calculated by Student's t-test.
