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Sequence of HC-toxin synthetase (HTS or Hts1), the product of HTS1. The four 'modules' are indicated in yellow. HTS1 was cloned using antibodies raised against a purified fragment of the protein, and by using oligonucleotides synthesized on the basis of  peptide sequences from the purified protein. Cloning was done by Jean-Alain Pocard, John Scott-Craig, and Dan Panaccione. The gene was sequenced (manually!) by John and Dan.

The role of HTS1 in HC-toxin biosynthesis was confirmed by targeted gene knockouts. All naturally occuring toxin-producing isolates of C. carbonum have two functional copies of this gene, and both must be disrupted to get a loss of virulence (see map of TOX2).

HTS1 contains no introns.

Hts1 contains four "modules" or "domains" (shown in yellow), one for amino acid. Each module activates its particular amino acid, forming an amino acyl-AMP derivative and subsequent covalent attachment to pantotheine. The order of the modules reflects the amino acid sequence of HC-toxin: module 1 activates L-proline, module 2 activates L-alanine, module 3 activates D-alanine, and module 4 activates L-Aeo.

Hts1 has one 'epimerization' motif, between modules 1 and 2, for converting L-Pro to D-Pro. The D-Ala in HC-toxin is made by a specific alanine racemase,  the product of TOXG. 

References:
Panaccione et al. (1992)  PNAS 89:6590-6594.
Scott-Craig et al. (1992) J. Biol. Chem. 67:26044-26049.
Cheng and Walton (2000) J. Biol. Chem. 275:4906-5004.