Sequence and peptide map of guinea pig aquaporin 0.
OBJECTIVE
Hyperbaric oxygen-treated guinea pigs serve as a useful animal model of nuclear cataract. To understand the structure and function of major intrinsic proteins in this model, the primary sequence and major posttranslational modifications to guinea pig aquaporin 0 (AQP0) were determined.


METHODS
The cDNA encoding guinea pig AQP0 was amplified by PCR, cloned and sequenced. After protein enrichment from guinea pig lens tissue, the protein sequence and the posttranslational modifications to AQP0 were determined by using combined chemical cleavage, trypsin and pepsin digestion with matrix assisted laser desorption/ionization mass spectrometry or capillary liquid chromatography tandem mass spectrometry.


RESULTS
The primary structure of AQP0 was determined from the DNA sequence and the translated sequence confirmed by mass spectrometry. Serine 235 was identified to be the major phosphorylation site.


CONCLUSIONS
Significant sequence homology was observed between species including putative regulatory sites of phosphorylation and pH regulation. These data form a foundation of information from which to begin assessing posttranslational modifications in cataract models.
Han
Jun
J
Department of Cell and Molecular Pharmacology, Medical University of South Carolina, Charleston, SC 29425, USA.
Little
Mark
M
David
Larry L
LL
Giblin
Frank J
FJ
Schey
Kevin L
KL
D000595:Amino Acid Sequence; D000818:Animals; D020346:Aquaporins; D001483:Base Sequence; D002853:Chromatography, Liquid; D005136:Eye Proteins / Q000737:chemistry* / Q000235:genetics; D006168:Guinea Pigs; D006863:Hydrogen-Ion Concentration; D007908:Lens, Crystalline / Q000737:chemistry*; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D010446:Peptide Fragments; D010449:Peptide Mapping; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D011499:Protein Processing, Post-Translational; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013329:Structure-Activity Relationship
D020346:Aquaporins; D005136:Eye Proteins; D008562:Membrane Glycoproteins; D010446:Peptide Fragments; C030328:aquaporin 0