1Y4K

Lipoxygenase-1 (Soybean) at 100K, N694G Mutant

Summary for 1Y4K

• Entry DOI: 10.2210/pdb1y4k/pdb
• Related: 1F8N 1FGM
• Descriptor: Seed lipoxygenase-1, FE (II) ION, ACETATE ION, ... (5 entities in total)
• Functional Keywords: lipoxygenase, dioxygenase, metalloprotein, fatty acids, iron, oxidoreductase
• Biological source: Glycine max (soybean)
• Cellular location: Cytoplasm: P08170
• Total number of polymer chains: 1
• Total formula weight: 94659.08

Authors

Chruszcz, M.,Segraves, E.,Holman, T.R.,Minor, W. (deposition date: 2004-12-01, release date: 2005-12-13, Last modification date: 2023-08-23)

Primary citation

Segraves, E.N.,Chruszcz, M.,Neidig, M.L.,Ruddat, V.,Zhou, J.,Wecksler, A.T.,Minor, W.,Solomon, E.I.,Holman, T.R.
Kinetic, spectroscopic, and structural investigations of the soybean lipoxygenase-1 first-coordination sphere mutant, Asn694Gly.
Biochemistry, 45:10233-10242, 2006

PubMed Abstract: In wild-type soybean LO-1 (WT sLO-1), Asn694 is a weak sixth ligand that is thought to be critical for enzymatic catalysis. In this investigation, N694G sLO-1 was studied to probe its contribution at this sixth ligand position to the kinetic and spectroscopic properties. The k(cat) value of N694G is approximately 230 times lower than that of WT sLO-1 at 25 degrees C, which can be partially explained by a lowered reduction potential of the iron as seen as a shift in the visible ligand-to-metal charge-transfer band (lambda(max) = 410 nm for N694G and lambda(max) = 425 nm for WT sLO-1). This conclusion was supported by a faster rate of oxidation of N694G by the product than that of WT sLO-1 (k(2) = 606 s(-)(1) for N694G and k(2) = 349 s(-)(1) for WT sLO-1). These results suggest a stronger ligand at the active site iron than the native Asn694, which is confirmed to be a water bound to the Fe(II) in the crystal structure. This produces a six-coordinate circular dichroism/magnetic circular dichroism (CD/MCD) spectra for ferrous N694G and an intermediate rhombic electron paramagnetic resonance (EPR) signal for ferric N694G. The EPR spectrum and its pH dependence suggest that the coordination environment of ferric N694G contains one hydroxide and one water. On the basis of both kinetic and structural factors, we propose that the Asn694 water-derived ligand would likely be a hydroxide and the active site, water-derived ligand a water in the ferric state, hence lowering the reaction rate of N694G more than would be expected from the lowered reduction potential alone.

PubMed: 16922498
DOI: 10.1021/bi060577e

Experimental method

X-RAY DIFFRACTION (1.95 Å)