Oligomerization of the Amide Sensor Protein AmiC by X-ray and Neutron Scattering and Molecular Modeling

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• 作者：Dean Chamberlain ; Bernard P. O'Hara ; Stuart A. Wilson ; Laurence H. Pearl ; and Stephen J. Perkins
• 刊名：Biochemistry
• 出版年：1997
• 出版时间：July 1, 1997
• 年：1997
• 卷：36
• 期：26
• 页码：8020 - 8029
• 全文大小：193K
• 年卷期：v.36,no.26(July 1, 1997)
• ISSN：1520-4995

文摘

AmiC is the negative regulator of the amidase operonwhich is involved in amide metabolismin the cytosol of Pseudomonas aeruginosa. Crystalstructures show that AmiC contains two large domainsthat are very similar to the periplasmic leucine-isoleucine-valinebinding protein (LivJ) of Escherichiacoli. Synchrotron X-ray and neutron (in 100%²H₂O buffer) scattering data were obtained forAmiC inthe presence of its substrate acetamide and its anti-inducer butyramidewhich binds more weakly to AmiCthan acetamide. Guinier analyses to obtain radius of gyrationR_G and molecular weightM_r values showedthat AmiC formed trimers whose formation was favored in the presence ofacetamide and which exhibitedconcentration-dependent properties at concentrations between 0.4 and 2mg/mL. Above 2 mg/mL, wheretrimers predominated, the R_G data were identicalwithin 0.05 nm for AmiC-acetamide and AmiC-butyramide with mean X-ray and neutron R_G valuesof 3.35 and 3.28 nm, respectively. Scattering curvefits constrained by the crystal structure of AmiC-acetamide wereevaluated in order to describe a modelfor trimeric AmiC. A translational search of parallel alignmentsof three monomers to form a symmetricAmiC homotrimer gave a good X-ray curve fit. Combinations ofcalculated curves for monomeric, dimeric,trimeric, and tetrameric AmiC as seen in the crystal structure of AmiCgave reasonable but weaker X-raycurve fits which did not favor the existence of tetrameric AmiC.It is concluded that AmiC exhibitsnovel ligand-dependent oligomerization properties in solution whenthese are compared to other membersof the periplasmic binding protein superfamily, where AmiC exists inmonomeric and trimeric forms, theproportions of which depend on the presence of acetamide orbutyramide.