Structure, function and dynamics in acyl carrier proteins.

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Carrier proteins are essential four-helix bundles that bind metabolites, including fatty acids and polyketides, crucial for synthesizing pharmaceutically important compounds like antibiotics.
• Acyl carrier proteins (ACPs) exist as single domains in multi-domain polypeptides (Type I ACPs) or as part of multiprotein complexes (Type II).

Purpose of the Study:

• To investigate the substrate-binding mechanisms of ACP2 and ACP3, domains of the type I trans-AT polyketide synthase MmpA involved in mupirocin antibiotic biosynthesis.
• To explore how specific amino acid residues influence the formation of substrate-binding pockets in ACPs.

Main Methods:

• Molecular dynamics simulations were performed on apo, holo, and acyl forms of ACP2 and ACP3.
• Site-directed mutagenesis was used to alter key residues in ACP3 and an E. coli fatty acid synthase ACP.

Main Results:

• Wild-type ACP2 and ACP3 exhibit a substrate-binding surface-groove where exposed polar groups interact with the solvent.
• Bulky hydrophobic residues in the conserved GXDS motif and helix III of Type I and Type II polyketide synthase ACPs prevent deep tunnel formation.
• Mutations in ACP3 (I61A L36A W44L) created a deep tunnel, fully burying a saturated substrate, unlike the wild type's surface groove.
• Mutations in an E. coli fatty acid synthase ACP also demonstrated a switch between deep tunnel and surface groove binding.

Conclusions:

• The structural characteristics of specific residues, particularly in the GXDS motif and helix III, dictate whether ACPs form surface-binding grooves or deep tunnels.
• Modifying a few key amino acid residues in ACPs can significantly alter their substrate-binding modes, offering insights into the engineering of biosynthetic pathways for novel compounds.