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Abstract Phosphopantetheinyl transferases (PPTases) play an essential role in primary and secondary metabolism. These enzymes facilitate the post-translational activation of acyl carrier proteins (ACPs) central to the biosynthesis of fatty acids and polyketides. Modulation of ACP-PPTase interactions is a promising approach to both increase access to desired molecular outputs and disrupt mechanisms associated with disease progression. However, such an approach requires understanding the molecular principles that govern ACP-PPTase interactions across diverse synthases. Through a multi-year, course-based undergraduate research experience (CURE), 17 ACPs representing a range of putative type II polyketide synthases, from actinobacterial and non-actinobacterial phyla, were evaluated as substrates for three PPTases (AcpS, Sfp, and vulPPT). The observed PPTase compatibility, sequence-level analyses, and predictive structural modelling suggest that ACP selectivity is driven by amino acids surrounding the conserved, modified serine on the ACP. We propose that vulPPT and Sfp are driven primarily by hydrophobic contacts, whereas AcpS may favor ACPs which exhibit high net-negative charge density, as well as a broad electronegative surface distribution. Furthermore, we report a plausible, hitherto unreported hydrophobic interaction between vulPPT and a conserved ACP crease, upstream of the invariant serine, which may facilitate docking. This work provides a catalog of compatible and incompatible ACP-PPTase partnerships, highlighting specific regions on the ACP and/or PPTase that show promise for future strategic engineering and inhibitor development efforts.
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The effect of divalent cations on the thermostability of type II polyketide synthase acyl carrier proteins
The successful engineering of biosynthetic pathways hinges on understanding the factors that influence acyl carrier protein (ACP) stability and function. The stability and structure of ACPs can be influenced by the presence of divalent cations, but how this relates to primary sequence remains poorly understood. As part of a course‐based undergraduate research experience, we investigated the thermostability of type II polyketide synthase (PKS) ACPs. We observed an approximate 40 °C range in the thermostability among the 14 ACPs studied, as well as an increase in stability (5–26 °C) of the ACPs in the presence of divalent cations. Distribution of charges in the helix II‐loop–helix III region was found to impact the enthalpy of denaturation. Taken together, our results reveal clues as to how the sequence of type II PKS ACPs relates to their structural stability, information that can be used to study how ACP sequence relates to function. © 2018 American Institute of Chemical EngineersAIChE J, 64: 4308–4318, 2018
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- Award ID(s):
- 1652424
- PAR ID:
- 10077237
- Author(s) / Creator(s):
- ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; more »
- Publisher / Repository:
- Wiley Blackwell (John Wiley & Sons)
- Date Published:
- Journal Name:
- AIChE Journal
- Volume:
- 64
- Issue:
- 12
- ISSN:
- 0001-1541
- Format(s):
- Medium: X Size: p. 4308-4318
- Size(s):
- p. 4308-4318
- Sponsoring Org:
- National Science Foundation
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