Probing the Phosphopantetheine Arm Conformations of Acyl Carrier Proteins Using Vibrational Spectroscopy

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• 作者：Matthew N. R. Johnson ; Casey H. Londergan ; Louise K. Charkoudian
• 刊名：Journal of the American Chemical Society
• 出版年：2014
• 出版时间：August 13, 2014
• 年：2014
• 卷：136
• 期：32
• 页码：11240-11243
• 全文大小：230K
• ISSN：1520-5126

文摘

Acyl carrier proteins (ACPs) are universal and highly conserved domains central to both fatty acid and polyketide biosynthesis. These proteins tether reactive acyl intermediates with a swinging 4鈥?phosphopantetheine (Ppant) arm and interact with a suite of catalytic partners during chain transport and elongation while stabilizing the growing chain throughout the biosynthetic pathway. The flexible nature of the Ppant arm and the transient nature of ACP鈥揺nzyme interactions impose a major obstacle to obtaining structural information relevant to understanding polyketide and fatty acid biosynthesis. To overcome this challenge, we installed a thiocyanate vibrational spectroscopic probe on the terminal thiol of the ACP Ppant arm. This site-specific probe successfully reported on the local environment of the Ppant arm of two ACPs previously characterized by solution NMR, and was used to determine the solution exposure of the Ppant arm of an ACP from 6-deoxyerythronolide B synthase (DEBS). Given the sensitivity of the probe鈥檚 CN stretching band to conformational distributions resolved on the picosecond time scale, this work lays a foundation for observing the dynamic action-related structural changes of ACPs using vibrational spectroscopy.