Abstract
The acyl-coenzyme A binding protein (ACBP) is a conserved regulator of several cell and phylogenetic functions, including lipid synthesis, energy metabolism, autophagy, and appetite stimulations. ACBP is a small four-helix bundle protein whose structure has been studied by NMR spectroscopy, but its thermal stability has not been studied by electron spin resonance (ESR). Here, we characterize the structural dynamics and determine site-specific local thermal stability of ACBP in apo versus holo (i.e., the palmitoyl-CoA bound form) states using circular dichroism, fluorescence, pulsed dipolar ESR spectroscopies, and the recently developed ESR-based peak-height method. We recorded ESR spectra of single-labeled apo and holo ACBP at temperatures from 300 to 355 K. The ESR spectra in the absorption mode (i.e., integrated ESR spectra) were analyzed to determine the onset of local disruption for individual sites during thermal denaturation. Using the ESR-based peak-height method, this study reveals a previously undescribed response of ACBP that the binding of palmitoyl-CoA to ACBP not only promotes the stability of the binding region (helices α1–α3) but also causes a distinct increase in the thermal stability of the interface between helices α1 and α4, a region distant from the binding site.
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The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.
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Acknowledgements
This work was supported by grants (108-2113-M-007-029 and 110-2731-M-007-001) from the Ministry of Science and Technology of Taiwan and the Frontier Research Center on Fundamental and Applied Sciences of Matters at NTHU. All the ESR measurements were conducted in the Research Instrument Center of Taiwan located at NTHU.
Funding
Ministry of Science and Technology, Taiwan, 108-2113-M-007-029, Yun-Wei Chiang, 110-2731-M-007-001,Yun-Wei Chiang
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Hung, CL., Lee, S.W. & Chiang, YW. Local Structural Stability of the Acyl-Coenzyme A Binding Protein by ESR Spectroscopy. Appl Magn Reson 54, 107–118 (2023). https://doi.org/10.1007/s00723-022-01476-w
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DOI: https://doi.org/10.1007/s00723-022-01476-w