Increasing nitrogenase catalytic efficiency for MgATP by changing serine 16 of its Fe protein to threonine: use of Mn2+ to show interaction of serine 16 with Mg2+

Protein Science : a Publication of the Protein Society
L C Seefeldt, L E Mortenson

Abstract

MgATP-binding and hydrolysis are an integral part of the nitrogenase catalytic mechanism. We are exploring the function of MgATP hydrolysis in this reaction by analyzing the properties of the Fe protein (FeP) component of Azotobacter vinelandii nitrogenase altered by site-directed mutagenesis. We have previously (Seefeldt, L.C., Morgan, T.V., Dean, D.R., & Mortenson, L.E., 1992, J. Biol. Chem. 267, 6680-6688) identified a region near the N-terminus of FeP that is involved in interaction with MgATP. This region of FeP is homologous to the well-known nucleotide-binding motif GXXXXGKS/T. In the present work, we examined the function of the four hydroxyl-containing amino acids immediately C-terminal to the conserved lysine 15 that is involved in interaction with the gamma-phosphate of MgATP. We have established, by altering independently Thr 17, Thr 18, and Thr 19 to alanine, that a hydroxyl-containing residue is not needed at these positions for FeP to function. In contrast, an hydroxyl-containing amino acid at position 16 was found to be critical for FeP function. When the strictly conserved Ser 16 was altered to Ala, Cys, Asp, or Gly, the FeP did not support N2 fixation when expressed in place of the wild-type FeP in A. vineland...Continue Reading

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Citations

Aug 5, 1997·Proceedings of the National Academy of Sciences of the United States of America·M K Sluis, S A Ensign
Nov 13, 2008·The Journal of Biological Chemistry·Jeffrey M BoydDiana M Downs
Sep 26, 2013·Proceedings of the National Academy of Sciences of the United States of America·Zhi-Yong YangLance C Seefeldt
Mar 21, 2008·Journal of Biological Inorganic Chemistry : JBIC : a Publication of the Society of Biological Inorganic Chemistry·Jan PetersenDavid J Lowe
Apr 4, 2008·Journal of Biological Inorganic Chemistry : JBIC : a Publication of the Society of Biological Inorganic Chemistry·Jan PetersenDavid J Lowe
Jan 26, 1996·The Journal of Biological Chemistry·L PulakatN Gavini
Nov 19, 2004·Journal of Biological Inorganic Chemistry : JBIC : a Publication of the Society of Biological Inorganic Chemistry·Se Bok JangJohn W Peters
Feb 29, 2000·The Journal of Biological Chemistry·M J Ryle, L C Seefeldt
Feb 28, 2003·Journal of Biological Inorganic Chemistry : JBIC : a Publication of the Society of Biological Inorganic Chemistry·Morten SørlieVernon D Parker
Oct 29, 1998·The Journal of Biological Chemistry·E H Bursey, B K Burgess
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Jan 18, 1994·Biochemistry·J Kim, D C Rees
Nov 7, 1996·Chemical Reviews·Barbara K. Burgess, David J. Lowe

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