Structural Characterization of an ACP from Thermotoga maritima: Insights into Hyperthermal Adaptation

Abstract

Thermotoga maritima, a deep-branching hyperthermophilic bacterium, expresses an extraordinarily stable Thermotoga maritima acyl carrier protein (Tm-ACP) that functions as a carrier in the fatty acid synthesis system at near-boiling aqueous environments. Here, to understand the hyperthermal adaptation of Tm-ACP, we investigated the structure and dynamics of Tm-ACP by nuclear magnetic resonance (NMR) spectroscopy. The melting temperature of Tm-ACP (101.4 • C) far exceeds that of other ACPs, owing to extensive ionic interactions and tight hydrophobic packing. The D59 residue, which replaces Pro/Ser of other ACPs, mediates ionic clustering between helices III and IV. This creates a wide pocket entrance to facilitate the accommodation of long acyl chains required for hyperthermal adaptation of the T. maritima cell membrane. Tm-ACP is revealed to be the first ACP that harbor an amide proton hyperprotected against hydrogen/deuterium exchange for I15. The hydrophobic interactions mediated by I15 appear to be the key driving forces of the global folding process of Tm-ACP. Our findings provide insights into the structural basis of the hyperthermal adaptation of ACP, which might have allowed T. maritima to survive in hot ancient oceans. 1 Thermotoga maritima ACP; 2 Pseudothermotoga thermarum ACP; 3 Thermus aquaticus ACP; 4 Enterococcus faecalis ACP; 5 Brucella melitenis ACP; 6 Escherichia coli ACP; 7 Vibrio harveyi ACP; 8 The isoelectric points (pI) of bacterial ACPs were calculated by ProtParam tool (https://web.expasy.org/protparam) [35].

Deciphering the Binding Interactions between Acinetobacter baumannii ACP and β-ketoacyl ACP Synthase III to Improve Antibiotic Targeting Using NMR Spectroscopy

“…This Leu in EcACP is reportedly important to open up a path into the binding cavity of ACP [ 20 ]. Interestingly, hyperthermophilic Thermotoga maritima ACP has phenylalanine at this position [ 32 ]. In this study, we investigated the solution structure of AbACP and the role of these residues in the structure and thermal stability of AbACP in comparison with other ACPs.…”

“…In AbACP, the mutation of F47 with Ala resulted in a dramatic decrease in thermal stability, confirming the importance of F47 in hydrophobic packing inside the cavity. F47 at the center of the α 2 -helix in AbACP is not conserved in most ACPs; however, it is observed as F50 in Thermotoga maritima ACP (TmACP) [ 32 ]. In our previous report, the substitution of Ala for F50 in TmACP was found to cause a decrease in the melting temperature from 100.4 to 90.7 °C.…”

“…In our previous report, the substitution of Ala for F50 in TmACP was found to cause a decrease in the melting temperature from 100.4 to 90.7 °C. In hyperthermostable TmACP, extremely tight hydrophobic packing occurs, as well as extensive ionic clusters on the surface of the protein [ 32 ]; thus, although a single F50A mutation was not capable of causing dramatic changes, it did significantly impact the thermal stability. In fact, in mesophilic AbACP, a single F47A mutation caused a 30-degree reduction in thermal stability, implying that F47 is the key residue for tight hydrophobic packing in AbACP.…”

“…In particular, the α 2 -helix of ACPs, referred to as the “recognition α 2 -helix”, contains acidic residues and is important for interactions with FAS enzymes [ 13 , 25 ]. NMR spectroscopy has been successfully applied for the identification of the resolution structures of type II ACPs in Escherichia coli (EcACP) [ 26 , 27 , 28 ], Vibrio harveyi (VhACP) [ 29 ], Borrelia burgdorferi (BbACP) [ 30 ], Enterococcus faecalis (EfACP) [ 31 ], and Thermotoga maritima (TmACP) [ 32 ].…”

Special Issue “Selected Papers from the 8th Asia-Pacific NMR (APNMR) Symposium: Recent Advances in NMR Spectroscopy”

“…The Cheng group makes a significant step forward in understanding the mechanism of action of antimicrobial peptide P113 interactions with a live fungal strain of Candida [ 5 ]. Yangmee Kim and coworkers report an NMR-derived 3D structure of an extremely thermostable acyl carrier protein from bacteria T. maritima [ 6 ]. The structural analysis revealed a potential mechanism for the thermostability of the protein.…”