Biophysical and structural studies reveal marginal stability of a crucial hydrocarbon biosynthetic enzyme acyl ACP reductase.

Scientific Reports
Ashima SharmaSyed Shams Yazdani

Abstract

Acyl-ACP reductase (AAR) is one of the two key cyanobacterial enzymes along with aldehyde deformylating oxygenase (ADO) involved in the synthesis of long-chain alkanes, a drop-in biofuel. The enzyme is prone to aggregation when expressed in Escherichia coli, leading to varying alkane levels. The present work attempts to investigate the crucial structural aspects of AAR protein associated with its stability and folding. Characterization by dynamic light scattering experiment and intact mass spectrometry revealed that recombinantly expressed AAR in E. coli existed in multiple-sized protein particles due to diverse lipidation. Interestingly, while thermal- and urea-based denaturation of AAR showed 2-state unfolding transition in circular dichroism and intrinsic fluorescent spectroscopy, the unfolding process of AAR was a 3-state pathway in GdnHCl solution suggesting that the protein milieu plays a significant role in dictating its folding. Apparent standard free energy [Formula: see text] of ~ 4.5 kcal/mol for the steady-state unfolding of AAR indicated borderline stability of the protein. Based on these evidences, we propose that the marginal stability of AAR are plausible contributing reasons for aggregation propensity and hence...Continue Reading

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Citations

Nov 1, 2021·Journal of Industrial Microbiology & Biotechnology·Humaira Parveen, Syed Shams Yazdani

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Software Mentioned

AMBER18
Malvern Zetasizer
LipidSearch
ANTECHAMBER
Scientific Biopharma Finder
AmberTools18
UCSF Chimera
CPPTRAJ
SigmaPlot
Thermo

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