Probing the interactions of an acyl carrier protein domain from the 6‐deoxyerythronolide B synthase

Abstract

The assembly-line architecture of polyketide synthases (PKSs) provides an opportunity to rationally reprogram polyketide biosynthetic pathways to produce novel antibiotics. A fundamental challenge toward this goal is to identify the factors that control the unidirectional channeling of reactive biosynthetic intermediates through these enzymatic assembly lines. Within the catalytic cycle of every PKS module, the acyl carrier protein (ACP) first collaborates with the ketosynthase (KS) domain of the paired subunit in its own homodimeric module so as to elongate the growing polyketide chain and then with the KS domain of the next module to translocate the newly elongated polyketide chain. Using NMR spectroscopy, we investigated the features of a structurally characterized ACP domain of the 6-deoxyerythronolide B synthase that contribute to its association with its KS translocation partner. Not only were we able to visualize selective protein-protein interactions between the two partners, but also we detected a significant influence of the acyl chain substrate on this interaction. A novel reagent, CF 3 -S-ACP, was developed as a 19 F NMR spectroscopic probe of protein-protein interactions. The implications of our findings for understanding intermodular chain translocation are discussed.

Structural analysis of protein–protein interactions in type I polyketide synthases

“…The ACP4 interacts with KS5-AT5 linker mainly by electrostatic interaction between E23 on ACP4 helix I and R551 on KS5-AT5 linker. This proposed interaction site between ACP n-1 and KS n is supported by the NMR titration experiments using labeled ACP2 and unlabeled KS3-AT3, in which 1 H- 15 N HSQC signals for some residues in helix I disappeared in the presence of the KS3-AT3 didomain (Charkoudian et al, 2011). …”

S‐Trifluoromethylation of Thiols by Hypervalent Iodine Reagents: A Joint Experimental and Computational Study

“…[12,[18][19][20] Interestingly,h owever,t he selectivity observed for the desired transformation is heavily dependento nt he reaction temperature (Figure1). This reaction, beside ab road substrate scope, features good solvent compatibility.…”

“…[17] Indeed, reagents such as 1 are well suited for late-stage trifluoromethylation reactions,i np articular of thiols. [20] Although the chemistry of iodanes is well established,o nly recent computational studies revealed the mechanistic versatility of l 3 -iodanes, reminiscent of that of transition metals. [20] Although the chemistry of iodanes is well established,o nly recent computational studies revealed the mechanistic versatility of l 3 -iodanes, reminiscent of that of transition metals.…”

Irreversible Cysteine‐Selective Protein Labeling Employing Modular Electrophilic Tetrafluoroethylation Reagents

“…Trifluoromethylation reagents based on hypervalent iodine ( 1 and 2 , Figure ) are known to transfer the trifluoromethyl group to various S‐, P‐, O‐, N‐, and C‐nucleophilic functionalities . Thiols, in particular, exhibit excellent reactivity and selectivity in such fluoroalkylation reactions, which has been demonstrated by selective S‐trifluoromethylation of, for example, coenzyme A and Sandostatin using 1 and 2 .…”