Water permeability of C-terminally truncated aquaporin 0 (AQP0 1-243) observed in the aging human lens.
OBJECTIVE
To first assess the distribution of posttranslationally truncated products of aquaporin 0 (AQP0) in dissected sections of a normal human lens and to determine the effect of backbone cleavage on the water permeability of AQP0.


METHODS
A 27-year-old lens was concentrically dissected into six sections. Membrane protein was isolated from each section and cleaved with cyanogen bromide, and the peptides were separated and analyzed by reverse-phase (RP)-HPLC-mass spectrometry (MS). The sites of posttranslational AQP0 C-terminal truncation were determined by mass spectrometry. Truncated forms of AQP0 were expressed in a Xenopus laevis oocyte system, and the effect of truncation on AQP0 water permeability was assessed in an oocyte osmotic swelling assay.


RESULTS
The extent of truncation at many sites within the C terminus increased with fiber cell age, and the effects of truncations after residues 234, 238, and 243 on AQP0 water permeability were examined. Truncation after residue 243 resulted in an approximate 15% decrease in permeability compared with the full-length protein, AQP0 1-263. However, rather than a direct effect on water transport, analysis of surface protein expression indicated that the decrease in permeability was a result of less efficient protein trafficking to the oocyte surface and that the permeabilities of full-length and 1-243 AQP0 were indistinguishable. Further, C-terminal truncation of AQP0 to 1-234 and 1-238, completely impaired trafficking into the plasma membrane, precluding the measurement of permeability.


CONCLUSIONS
These data provide evidence that loss of 20 amino acids from the C terminus may not directly affect the ability of AQP0 to transport water.
Department of Pharmacology, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
Lauren E
LE
Ball
Mark
M
Little
Mark W
MW
Nowak
Donita L
DL
Garland
Rosalie K
RK
Crouch
Kevin L
KL
Schey
D000328:Adult; D000375:Aging / Q000502:physiology*; D000596:Amino Acids; D000818:Animals; D020346:Aquaporins; D001693:Biological Transport, Active; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D005136:Eye Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D015870:Gene Expression; D006801:Humans; D007908:Lens, Crystalline / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000378:metabolism; D010539:Permeability; D017384:Sequence Deletion; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013329:Structure-Activity Relationship; D014867:Water / Q000378:metabolism*; D014982:Xenopus laevis
D000596:Amino Acids; D020346:Aquaporins; D005136:Eye Proteins; D008562:Membrane Glycoproteins; C030328:aquaporin 0; D014867:Water