Isolation and characterization of hydrophobic polypeptides in human bile.
Polypeptides were isolated from human bile by extraction with chloroform/methanol, followed by reversed-phase chromatography in methanol/ethylene chloride and gel filtration in chloroform/methanol. Peptides were characterized by SDS/PAGE, sequence analysis and matrix-assisted laser desorption ionization/time-of-flight mass spectrometry. This identified haemoglobin alpha chain, ATP synthase lipid-binding protein subunit 9, an N-terminal fragment of mac25/insulin-like growth factor-binding protein 7 and an internal fragment of monocyte differentiation antigen CD14, all not described previously in bile. In addition, alpha1-antitrypsin, known in bile from previous work, was also identified. The hydrophobic character of haemoglobin alpha chain is not apparent from its amino acid sequence, but the other polypeptides all have major hydrophobic segments. These results show that several proteins are removed upon organic solvent extraction used for delipidation during the preparation of samples for proteome analysis. Several of the polypeptides found are unexpectedly present in bile, suggesting that specific excretion mechanisms may be involved.
Stark
M
M
Department of Medical Biochemistry, Karolinska Institutet, Stockholm, Sweden. Margareta, Stark@mbb.ki.se
Jörnvall
H
H
Johansson
J
J
D000595:Amino Acid Sequence; D001646:Bile / Q000737:chemistry*; D002352:Carrier Proteins / Q000737:chemistry; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002784:Cholesterol / Q000378:metabolism; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003460:Crystallization; D003911:Dextrans; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins; D005704:Gallbladder / Q000378:metabolism; D006443:Hemoglobin A2 / Q000737:chemistry; D006801:Humans; D018969:Insulin-Like Growth Factor Binding Proteins; D018950:Lipopolysaccharide Receptors / Q000737:chemistry; D025261:Mitochondrial Proton-Translocating ATPases; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification*; D006180:Proton-Translocating ATPases / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D012997:Solvents; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D000515:alpha 1-Antitrypsin / Q000737:chemistry
D002352:Carrier Proteins; D003911:Dextrans; D005656:Fungal Proteins; D018969:Insulin-Like Growth Factor Binding Proteins; D018950:Lipopolysaccharide Receptors; D010446:Peptide Fragments; D010455:Peptides; D012997:Solvents; D000515:alpha 1-Antitrypsin; C102529:insulin-like growth factor binding protein-related protein 1; D006443:Hemoglobin A2; C040462:sephadex, 2-hydroxypropyl ether; D002784:Cholesterol; D025261:Mitochondrial Proton-Translocating ATPases; D006180:Proton-Translocating ATPases