Glomerular charge selectivity for anionic and neutral horseradish peroxidase.
Studies in isolated perfused rat kidney have demonstrated that it exhibits apparently normal charge selectivity and tubular uptake of anionic horseradish peroxidase (aHRP; pI < 4.0) and neutral horseradish peroxidase (nHRP; pI = 7.5) when these proteins are measured for their enzyme activity. The absolute fractional clearance values for aHRP and nHRP were 0.006 +/- 0.002 and 0.041 +/- 0.007, respectively. It is evident, however, that the enzyme assay for horseradish peroxidase severely underestimates the quantity of protein in urine as compared to measurement of its tritium labeled form through radioactivity. Fractional clearances estimated by radioactivity and corrected for tubular reabsorption for [3H]aHRP and [3H]nHRP were 0.040 +/- 0.029 and 0.099 +/- 0.043, respectively, compared to those estimated by enzyme activity which were 0.012 +/- 0.004 and 0.070 +/- 0.037, respectively. While charge selectivity between the anionic and neutral forms of HRP was still evident, albeit significantly reduced, the major feature of this type of analysis is that the clearance of the aHRP protein is significantly increased compared to that determined by enzyme assay. This difference correlates with the observation that the aHRP protein is markedly degraded (61 to 65%), as determined by gel chromatography, during filtration. Similar degradation was seen in urine fractions collected after the aHRP protein was administered in vivo. Degradation also occurred for the nHRP protein in both the perfused kidney and in vivo but to a far lesser extent (approximately 14 to 21%). These studies demonstrate that the anionic form of HRP was preferentially degraded during filtration and that charge selectivity for differently charged proteins is not as marked as originally thought.
Osicka
T M
TM
Biochemistry Department, Monash University, Clayton, Victoria, Australia.
Comper
W D
WD
D000818:Animals; D000838:Anions / Q000378:metabolism*; D004594:Electrophysiology; D006735:Horseradish Peroxidase / Q000493:pharmacokinetics* / Q000652:urine; D066298:In Vitro Techniques; D007668:Kidney / Q000378:metabolism; D007678:Kidney Glomerulus / Q000502:physiology*; D008297:Male; D051381:Rats; D011920:Rats, Inbred WF; D017207:Rats, Sprague-Dawley
D000838:Anions; D006735:Horseradish Peroxidase