Functional expression and properties of Sec14p-like protein with molecular mass 45 kD from rat olfactory epithelium.
cDNA of Sec14p-like water-soluble protein with molecular mass 45 kD from rat olfactory epithelium was expressed in Escherichia coli Rosetta cells. The expression product was purified by a two-step chromatographic procedure on DEAE-Sepharose and Sephacryl S-200. The identity of structural and functional characteristics of the recombinant and native proteins was demonstrated by CD, mass spectrometry, and Western blotting. Using several lipids immobilized on nitrocellulose membranes, it was shown that phosphatidylinositol-3,4,5-triphosphate is the specific ligand for the studied protein.
Radchenko
V V
VV
Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, 117997, Russia. radchenko@mail.ibch.ru
Merkulova
M I
MI
Shuvaeva
T M
TM
Simonova
T N
TN
Bondar
A A
AA
Lipkin
V M
VM
D000818:Animals; D015153:Blotting, Western; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism; D008024:Ligands; D013058:Mass Spectrometry; D008970:Molecular Weight; D009831:Olfactory Mucosa / Q000378:metabolism*; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification
D002352:Carrier Proteins; D008024:Ligands; D018129:Phosphatidylinositol Phosphates; D011994:Recombinant Proteins; C506000:Sec14p-like phosphoinositide-binding p45 protein, rat; C060974:phosphatidylinositol 3,4,5-triphosphate