All polyketide synthases, fatty acid synthases, and non-ribosomal peptide synthetases
require posttranslational modification of their constituent acyl carrier protein domain(s)
to become catalytically active. The inactive apoproteins are converted to their active
holo-forms by posttranslational transfer of the 4'-phosphopantetheinyl (P-pant) moiety
of coenzyme A to the sidechain hydroxyl of a conserved serine residue in each acyl
carrier protein domain. The first P-pant transferase to be cloned and characterized
was the recently reported Escherichia coli enzyme ACPS, responsible for apo to holo
conversion of fatty acid synthase. Surprisingly, initial searches of sequence databases
did not reveal any proteins with significant peptide sequence similarity with ACPS.
Through refinement of sequence alignments that indicated low level similarity with
the ACPS peptide sequence, we identified two consensus motifs shared among several
potential ACPS homologs. This has led to the identification of a large family of proteins
having 12-22 % similarity with ACPS, which are putative P-pant transferases. Three
of these proteins, E. coli EntD and o195, and B. subtilis Sfp, have been overproduced,
purified and found to have P-pant transferase activity, confirming that the observed
low level of sequence homology correctly predicted catalytic function. Three P-pant
transferases are now known to be present in E. coli (ACPS, EntD and o195); ACPS and
EntD are specific for the activation of fatty acid synthase and enterobactin synthetase,
respectively. The apo-protein substrate for o195 has not yet been identified. Sfp
is responsible for the activation of the surfactin synthetase.
The specificity of ACPS and EntD for distinct P-pant-requiring enzymes suggests that
each P-pant-requiring synthase has its own partner enzyme responsible for apo to holo
activation of its acyl carrier domains. This is the first direct evidence that in
organisms containing multiple P-pant-requiring pathways, each pathway has its own
posttranslational modifying activity.