title: Mass spectral determination of phosphopantetheinylation specificity for carrier proteins inMycobacterium tuberculosis.
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 inMycobacterium 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.
authors.affiliation: Maurice Wilkins Centre for Molecular Biodiscovery and School of Biological Sciences The University of Auckland New Zealand; Present address: W. M. Keck Structural Biology Laboratory Cold Spring Harbor Laboratory 1 Bungtown Road Cold Spring Harbor NY 11724 USA.
authors.forename: James
authors.initials: J
authors.lastname: Jung
authors.affiliation: Maurice Wilkins Centre for Molecular Biodiscovery and School of Biological Sciences The University of Auckland New Zealand.
authors.forename: Ghader
authors.identifier: 0000-0002-5092-3749
authors.initials: G
authors.lastname: Bashiri
authors.affiliation: Maurice Wilkins Centre for Molecular Biodiscovery and School of Biological Sciences The University of Auckland New Zealand.
authors.forename: Jodie M
authors.initials: JM
authors.lastname: Johnston
authors.affiliation: Maurice Wilkins Centre for Molecular Biodiscovery and School of Biological Sciences The University of Auckland New Zealand.
authors.forename: Edward N
authors.initials: EN
authors.lastname: Baker
keywords: Mycobacterium tuberculosis; carrier proteins; mass spectrometry; mutagenesis; phosphopantetheinyl transferases