Abstract
Soybean lipoxygenase-1 (LOX-1) is used widely as a model for studying the structural and functional properties of the homologous family of lipoxygenases. The crystallographic structure revealed that LOX-1 is organized in a β-sheet N-terminal domain and a larger, mostly helical, C-terminal domain. Here, we describe the overall structural characterization of native unliganded LOX-1 in solution, using small angle X-ray scattering (SAXS). We show that the scattering pattern of the unliganded enzyme in solution does not display any significant difference compared with that calculated from the crystal structure, and that models of the overall shape of the protein calculated ab initio from the SAXS pattern provide a close envelope to the crystal structure. These data, demonstrating that LOX-1 has a compact structure also in solution, rule out any major motional flexibility of the LOX-1 molecule in aqueous solutions. In addition we show that eicosatetraynoic acid, an irreversible inhibitor of lipoxygenase used to mimic the effect of substrate binding, does not alter the overall conformation of LOX-1 nor its ability to bind to membranes. In contrast, the addition of glycerol (to 5%, v/v) causes an increase in the binding of the enzyme to membranes without altering its catalytic efficiency towards linoleic acid nor its SAXS pattern, suggesting that the global conformation of the enzyme is unaffected. Therefore, the compact structure determined in the crystal appears to be essentially preserved in these various solution conditions. During the preparation of this article, a paper by M. Hammel and co-workers showed instead a sharp difference between crystal and solution conformations of rabbit 15-LOX-1. The possible cause of this difference might be the presence of oligomers in the rabbit lipoxygenase preparations.
| Original language | English |
|---|---|
| Pages (from-to) | 143-152 |
| Number of pages | 10 |
| Journal | Journal of Molecular Biology |
| Volume | 349 |
| Issue number | 1 |
| DOIs | |
| Publication status | Published - May 27 2005 |
Fingerprint
Keywords
- Calcium
- Catalysis
- Inhibitors
- Membranes
- Soybean lipoxygenase-1
ASJC Scopus subject areas
- Virology
Cite this
Structural stability of soybean lipoxygenase-1 in solution as probed by small angle x-ray scattering. / Dainese, Enrico; Sabatucci, Annalaura; Van Zadelhoff, Guus; Angelucci, Clotilde Beatrice; Vachette, Patrice; Veldink, Gerrit A.; Agrò, Alessandro Finazzi; Maccarrone, Mauro.
In: Journal of Molecular Biology, Vol. 349, No. 1, 27.05.2005, p. 143-152.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Structural stability of soybean lipoxygenase-1 in solution as probed by small angle x-ray scattering
AU - Dainese, Enrico
AU - Sabatucci, Annalaura
AU - Van Zadelhoff, Guus
AU - Angelucci, Clotilde Beatrice
AU - Vachette, Patrice
AU - Veldink, Gerrit A.
AU - Agrò, Alessandro Finazzi
AU - Maccarrone, Mauro
PY - 2005/5/27
Y1 - 2005/5/27
N2 - Soybean lipoxygenase-1 (LOX-1) is used widely as a model for studying the structural and functional properties of the homologous family of lipoxygenases. The crystallographic structure revealed that LOX-1 is organized in a β-sheet N-terminal domain and a larger, mostly helical, C-terminal domain. Here, we describe the overall structural characterization of native unliganded LOX-1 in solution, using small angle X-ray scattering (SAXS). We show that the scattering pattern of the unliganded enzyme in solution does not display any significant difference compared with that calculated from the crystal structure, and that models of the overall shape of the protein calculated ab initio from the SAXS pattern provide a close envelope to the crystal structure. These data, demonstrating that LOX-1 has a compact structure also in solution, rule out any major motional flexibility of the LOX-1 molecule in aqueous solutions. In addition we show that eicosatetraynoic acid, an irreversible inhibitor of lipoxygenase used to mimic the effect of substrate binding, does not alter the overall conformation of LOX-1 nor its ability to bind to membranes. In contrast, the addition of glycerol (to 5%, v/v) causes an increase in the binding of the enzyme to membranes without altering its catalytic efficiency towards linoleic acid nor its SAXS pattern, suggesting that the global conformation of the enzyme is unaffected. Therefore, the compact structure determined in the crystal appears to be essentially preserved in these various solution conditions. During the preparation of this article, a paper by M. Hammel and co-workers showed instead a sharp difference between crystal and solution conformations of rabbit 15-LOX-1. The possible cause of this difference might be the presence of oligomers in the rabbit lipoxygenase preparations.
AB - Soybean lipoxygenase-1 (LOX-1) is used widely as a model for studying the structural and functional properties of the homologous family of lipoxygenases. The crystallographic structure revealed that LOX-1 is organized in a β-sheet N-terminal domain and a larger, mostly helical, C-terminal domain. Here, we describe the overall structural characterization of native unliganded LOX-1 in solution, using small angle X-ray scattering (SAXS). We show that the scattering pattern of the unliganded enzyme in solution does not display any significant difference compared with that calculated from the crystal structure, and that models of the overall shape of the protein calculated ab initio from the SAXS pattern provide a close envelope to the crystal structure. These data, demonstrating that LOX-1 has a compact structure also in solution, rule out any major motional flexibility of the LOX-1 molecule in aqueous solutions. In addition we show that eicosatetraynoic acid, an irreversible inhibitor of lipoxygenase used to mimic the effect of substrate binding, does not alter the overall conformation of LOX-1 nor its ability to bind to membranes. In contrast, the addition of glycerol (to 5%, v/v) causes an increase in the binding of the enzyme to membranes without altering its catalytic efficiency towards linoleic acid nor its SAXS pattern, suggesting that the global conformation of the enzyme is unaffected. Therefore, the compact structure determined in the crystal appears to be essentially preserved in these various solution conditions. During the preparation of this article, a paper by M. Hammel and co-workers showed instead a sharp difference between crystal and solution conformations of rabbit 15-LOX-1. The possible cause of this difference might be the presence of oligomers in the rabbit lipoxygenase preparations.
KW - Calcium
KW - Catalysis
KW - Inhibitors
KW - Membranes
KW - Soybean lipoxygenase-1
UR - http://www.scopus.com/inward/record.url?scp=18144375533&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=18144375533&partnerID=8YFLogxK
U2 - 10.1016/j.jmb.2005.03.027
DO - 10.1016/j.jmb.2005.03.027
M3 - Article
C2 - 15876374
AN - SCOPUS:18144375533
VL - 349
SP - 143
EP - 152
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
SN - 0022-2836
IS - 1
ER -