Figure 5
From: Regulation of homocysteine metabolism by Mycobacterium tuberculosis S-adenosylhomocysteine hydrolase
Analysis of phosphorylation sites on Mtb-SahH.
(A) Diagrammatic representation of phosphorylation sites at SahH-P identified by mass-spectrometry. Five Thr (Thr216, Thr219, Thr220, Thr221 and Thr257) and one Ser residues (Ser376) were identified and labeled above the respective domains of SahH. Domain I is shown in blue color and domain II is shown in white color. (B) and (C) Histograms depicting relative phosphorylation of SahH and its single or multiple Thr mutants, respectively. Purified SahH-WT and its mutants were analyzed by immunoblotting with anti-pThr antibodies. Percent relative phosphorylation was calculated considering the phosphorylation intensity of SahH-WT as 100%. Corresponding immunoblots are shown in Supplementary Fig. S7A and S7B. (D) Metabolic labeling of SahH-WT and SahH-T219A/T220A/T221A mutant. Histogram depicting densitometric analysis of autoradiograph of metabolically labeled proteins purified from E. coli. Percent relative phosphorylation is shown considering phosphorylation intensity of SahH-WT as 100%. Corresponding autoradiograph and coomassie-stained gel image is shown in Supplementary Fig. S8.
