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Structural and functional characterization of a 5,10‐methenyltetrahydrofolate synthetase from Mycoplasma pneumoniae (GI: 13508087)
Author(s) -
Chen Shengfeng,
Yakunin Alexander F.,
Proudfoot Michael,
Kim Rosalind,
Kim SungHou
Publication year - 2005
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.20591
Subject(s) - mycoplasma pneumoniae , apyrase , enzyme , chemistry , biochemistry , adenosine triphosphate , adenosine diphosphate , divalent , biology , platelet , medicine , platelet aggregation , immunology , pneumonia , organic chemistry
Mycoplasma pneumoniae 5,10‐methenyltetrahydrofolate synthetase [MTHFS; also known as 5‐formyltetrahydrofolate cycloligase; Enzyme Commission (EC) 6.3.3.2] belongs to a large cycloligase protein family with 97 sequence homologues from bacteria to human. To help define the molecular (biochemical and biophysical) function of the M. pneumoniae MTHFS, we have previously determined its crystal structure at 2.2 Å resolution (Chen et al., Proteins 2004;56:839–843). In this current study, activity assays confirmed the functionality of the recombinant protein, with K m = 165 μ M for 5‐formyltetrahydrofolate (5‐FTHF) and K m = 166 μ M for MgATP. The methenyltetrahydrofolate activity of M. pneumoniae MTHFS has a requirement for divalent metal ions with Mg 2+ being most effective, and an absolute requirement for nucleoside 5′‐triphosphates with adenosine triphosphate (ATP) being most effective. Crystallization in the presence of substrates (MgATP, with or without 5‐FTHF) produced the complex structures of the protein with adenosine diphosphate (ADP) and phosphate at 2.2 Å resolution; with ADP, phosphate, and 5‐FTHF at 2.5 Å resolution. These structures directly demonstrated that the role of Mg 2+ in the reaction is to form the ATPMg 2+ –enzyme complex. Proteins 2005. © 2005 Wiley‐Liss, Inc.