Hydrophobic probe binding of beta-lactoglobulin in the native and molten globule state induced by high pressure as affected by pH, KIO(3) and N-ethylmaleimide.
High hydrostatic pressure (HHP) at 500 MPa and 50 degrees C induces beta-LG into the molten globule state. Retinol, cis-parinaric acid (CPA), and 1-anilino-naphthalene-8-sulfonate (ANS) fluorescence from pH 2.5 to 10.5 in the presence of the native and molten globule states of beta-LG indicate that retinol binds to beta-LG in the calyx, CPA at the surface hydrophobic site, and ANS in multiple hydrophobic sites. HHP treatment results in a decrease of beta-LG affinity for retinol and CPA, suggesting conformational changes in the calyx and surface hydrophobic site of beta-LG during HHP treatment. beta-LG treated by HHP in the presence of N-ethylmaleimide (NEM) retains retinol affinity, suggesting that NEM protects the calyx conformation of beta-LG during HHP treatment. HHP treatment of beta-LG in the presence of KIO(3) exhibits a great decrease of CPA affinity compared to HHP-treated beta-LG in the absence of KIO(3), suggesting the formation of non-native disulfide bonding at the CPA binding site.
Yang
Jian
J
Department of Food Science and Human Nutrition, Washington State University, Pullman 99163-6376, USA.
Powers
Joseph R
JR
Clark
Stephanie
S
Dunker
A Keith
AK
Swanson
Barry G
BG
D000817:Anilino Naphthalenesulfonates / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005033:Ethylmaleimide / Q000494:pharmacology*; D005231:Fatty Acids, Unsaturated / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006874:Hydrostatic Pressure; D007452:Iodates / Q000494:pharmacology*; D007782:Lactoglobulins / Q000378:metabolism*; D017680:Potassium Compounds / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence; D014801:Vitamin A / Q000378:metabolism
D000817:Anilino Naphthalenesulfonates; D005231:Fatty Acids, Unsaturated; D007452:Iodates; D007782:Lactoglobulins; D017680:Potassium Compounds; D014801:Vitamin A; C027132:1-anilino-8-naphthalenesulfonate; C039693:potassium iodate; D005033:Ethylmaleimide; C013641:parinaric acid