Mass spectral determination of phosphopantetheinylation specificity for carrier proteins in Mycobacterium tuberculosis. Issue 12 (24th October 2016)

• Record Type: Journal Article
• Authors: Jung, James; Bashiri, Ghader; Johnston, Jodie M.; Baker, Edward N.
• Is Part Of: FEBS open bio. Volume 6:Issue 12(2016)
• Page Start: 1220
• Page End: 1226
• Publication Date: 2016-10-24
• Subjects: carrier proteins -- mass spectrometry -- mutagenesis -- Mycobacterium tuberculosis -- phosphopantetheinyl transferases; Molecular biology -- Periodicals; Cytology -- Periodicals; Life sciences -- Periodicals; Biological Science Disciplines -- Periodicals; Molecular Biology -- Periodicals; Cell Biology -- Periodicals; Cytology; Life sciences; Molecular biology; Periodicals; 572.805
• DOI: 10.1002/2211-5463.12140
• Languages: English
• ISSNs: 2211-5463

Abstract

Abstract : Phosphopantetheinyl transferases (PPTases) are key elements in the modular syntheses performed by multienzyme systems such as polyketide synthases. PPTases transfer phosphopantetheine derivatives from Coenzyme A to carrier proteins (CPs), thus orchestrating substrate supply. We describe an efficient mass spectrometry‐based protocol for determining CP specificity for a particular PPTase in organisms possessing several candidate PPTases. We show that the CPs MbtL and PpsC, both involved in synthesis of essential metabolites in Mycobacterium tuberculosis, are exclusively activated by the type 2 PPTase PptT and not the type 1 AcpS. The assay also enables conclusive identification of the reactive serine on each CP. Abstract : Using mass spectrometry, we show that the phosphopantetheinyl carrier proteins MbtL and PpsC, both involved in synthesis of essential metabolites in Mycobacterium tuberculosis, are activated only by the type 2 phosphopantetheinyl transferase PptT, and not the type 1 AcpS. This protocol can be used to determine the activation specificity of other carrier proteins.