title: Structure and function of Mycobacterium smegmatis 7-keto-8-aminopelargonic acid (KAPA) synthase.
abstract: The biotin biosynthesis pathway is an attractive target for development of novel drugs against mycobacterial pathogens, however there are as yet no suitable inhibitors that target this pathway in mycobacteria. 7-Keto-8-aminopelargonic acid synthase (KAPA synthase, BioF) is the enzyme which catalyzes the first committed step of the biotin synthesis pathway, but both its structure and function in mycobacteria remain unresolved. Here we present the crystal structure of Mycobacterium smegmatis BioF (MsBioF). The structure reveals an incomplete dimer, and the active site organization is similar to, but distinct from Escherichia coli 8-amino-7-oxononanoate synthase (EcAONS), the E. coli homologue of BioF. To investigate the influence of structural characteristics on the function of MsBioF, we deleted bioF in M. smegmatis and confirmed that BioF is required for growth in the absence of exogenous biotin. Based on structural and mutagenesis studies, we confirmed that pyridoxal 5'-phosphate (PLP) binding site residues His129, Lys235 and His200 are essential for MsBioF activity in vivo and residue Glu171 plays an important, but not essential role in MsBioF activity. The N-terminus (residues 1-37) is also essential for MsBioF activity in vivo. The structure and function of MsBioF reported here provides further insights for developing new anti-tuberculosis inhibitors aimed at the biotin synthesis pathway.
authors.affiliation: State Key Laboratory of Microbial Technology, School of Life Sciences, Shandong University, Jinan, Shandong 250100, China.
authors.forename: Shanghua
authors.initials: S
authors.lastname: Fan
authors.affiliation: National Laboratory of Biomacromolecules and CAS Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: De-Feng
authors.initials: DF
authors.lastname: Li
authors.affiliation: National Laboratory of Biomacromolecules and CAS Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Da-Cheng
authors.initials: DC
authors.lastname: Wang
authors.affiliation: National Laboratory of Biomacromolecules and CAS Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Joy
authors.initials: J
authors.lastname: Fleming
authors.affiliation: National Laboratory of Biomacromolecules and CAS Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Hongtai
authors.initials: H
authors.lastname: Zhang
authors.affiliation: National Laboratory of Biomacromolecules and CAS Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Center for Tuberculosis Control of Guangdong Province, Guangdong 510630, China.
authors.forename: Lin
authors.initials: L
authors.lastname: Zhou
authors.affiliation: The 4th People's Hospital, Foshan, Guangdong 528000, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhou
authors.affiliation: Center for Tuberculosis Control of Guangdong Province, Guangdong 510630, China.
authors.forename: Tao
authors.initials: T
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Microbial Technology, School of Life Sciences, Shandong University, Jinan, Shandong 250100, China. Electronic address: guanjun@sdu.edu.cn.
authors.forename: Guanjun
authors.initials: G
authors.lastname: Chen
authors.affiliation: National Laboratory of Biomacromolecules and CAS Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China. Electronic address: zhangxe@sun5.ibp.ac.cn.
authors.forename: Xian-En
authors.initials: XE
authors.lastname: Zhang
authors.affiliation: National Laboratory of Biomacromolecules and CAS Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China. Electronic address: blj@ibp.ac.cn.
authors.forename: Lijun
authors.initials: L
authors.lastname: Bi
mesh_terms: D000596:Amino Acids / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001710:Biotin / Q000378:metabolism; D020134:Catalytic Domain; D020102:Mycobacterium smegmatis / Q000201:enzymology* / Q000378:metabolism; D011732:Pyridoxal Phosphate / Q000378:metabolism
chemical_list: D000596:Amino Acids; D001426:Bacterial Proteins; C001502:7-keto-8-aminopelargonic acid; D011732:Pyridoxal Phosphate; D001710:Biotin
keywords: 7-Keto-8-aminopelargonic acid (KAPA) synthase; Active site; Biotin synthesis pathway; Crystal structure; Mycobacterium smegmatis