Afamin is a novel human vitamin E-binding glycoprotein characterization and in vitro expression.
Hydrophobic vitamins are transported in human plasma and extravascular fluids by carrier proteins. No specific protein has been described so far for vitamin E, which plays a crucial role in protecting against oxidative damage and disease. We report here the purification of a 75-kDa glycoprotein with vitamin E-binding properties by stepwise chromatography of lipoprotein-depleted human plasma and monitoring of vitamin E (alpha-tocopherol)-binding activity. Partial sequencing identified this protein as afamin, a previously described member of the albumin gene family with four or five potential N-glycosylation sites. Glycosylation analysis indicated that >90% of the glycans were sialylated biantennary complex structures. The vitamin E-binding properties were confirmed using recombinantly expressed afamin. Qualitative and quantitative analysis of plasma and extravascular fluids revealed an abundant presence of this protein not only in plasma (59.8+/-13.3 microg/mL) but also in extravascular fluids such as follicular (34.4+/-12.7 microg/mL) and cerebrospinal (0.28+/-0.16 microg/mL) fluids, suggesting potential roles for afamin in fertility and neuroprotection. Afamin is partly (13%) bound to plasma lipoproteins. Afamin and vitamin E concentrations significantly correlate in follicular and cerebrospinal fluids but not in plasma. The vitamin E association of afamin in follicular fluid was directly demonstrated by gel filtration chromatography and immunoprecipitation which complements the in vitro findings for purified native and recombinant afamin.
Jerkovic
Lidija
L
Division of Genetic Epidemiology, Department of Medical Genetics, Clinical and Molecular Pharmacology, Innsbruck Medical University, Schoepfstrasse 41, A-6020 Innsbruck, Austria.
Voegele
Andreas F
AF
Chwatal
Sabine
S
Kronenberg
Florian
F
Radcliffe
Catherine M
CM
Wormald
Mark R
MR
Lobentanz
Eva M
EM
Ezeh
Benjie
B
Eller
Patricia
P
Dejori
Norbert
N
Dieplinger
Benjamin
B
Lottspeich
Friedrich
F
Sattler
Wolfgang
W
Uhr
Manfred
M
Mechtler
Karl
K
Dwek
Raymond A
RA
Rudd
Pauline M
PM
Baier
Gottfried
G
Dieplinger
Hans
H
D001692:Biological Transport; D001826:Body Fluids / Q000737:chemistry; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002845:Chromatography; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D047468:Immunoprecipitation; D008074:Lipoproteins / Q000378:metabolism; D011485:Protein Binding; D012709:Serum Albumin / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D000075462:Serum Albumin, Human; D014810:Vitamin E / Q000378:metabolism*
C088246:AFM protein, human; D002352:Carrier Proteins; D006023:Glycoproteins; D008074:Lipoproteins; D012709:Serum Albumin; D014810:Vitamin E; D000075462:Serum Albumin, Human