Reversible cryoactivation of recombinant human prorenin
Cleavage of prorenin's prosegment causes irreversible formation of renin. In contrast, renin activity is reversibly exposed when prorenin is acidified to pH 3.3. Nonetheless, acidification of plasma results in irreversible activation of prorenin, because endogenous proteases cleave the prosegme...
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Veröffentlicht in: | The Journal of biological chemistry 1992-06, Vol.267 (17), p.11753-11759 |
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Zusammenfassung: | Cleavage of prorenin's prosegment causes irreversible formation of renin. In contrast, renin activity is reversibly exposed
when prorenin is acidified to pH 3.3. Nonetheless, acidification of plasma results in irreversible activation of prorenin,
because endogenous proteases cleave the prosegment of acid-activated prorenin. Chilling of plasma results in irreversible
cryoactivation of prorenin. In this study we investigated whether cryoactivation of purified prorenin is reversible. The intrinsic
renin activity of recombinant human prorenin was measured by an enzyme kinetic assay using partially purified human angiotensinogen
as substrate. Results are expressed as a percent (mean +/- S.E.) of the maximal activity exposed after limited proteolysis
by trypsin. The intrinsic renin activity of two pools (0.3 and 0.06 Goldblatt units/ml) was 1.5% +/- 0.3 and 1.2% +/- 0.6
at 37 degrees C. Activity increased to 19% +/- 0.3 and 26% +/- 0.5 after incubation at 0 degrees C and to 5.4% +/- 0.5 and
2.1% +/- 1.2 at room temperature. Cryoactivation did not occur in buffers containing more than 1 M NaCl. It took 8 min at
37 degrees C or 180 min at room temperature for cryoactivated prorenin to lose half of its intrinsic renin activity. It took
48 and 26 h, respectively, at 0 degree C for the two pools of prorenin at 37 degrees C to regain half of their maximum intrinsic
activity at 0 degrees C. A direct immunoradiometric assay that detects active renin but not prorenin was able to detect cryoactivated
prorenin. These results show that human prorenin can be reversibly cryoactivated in buffers of low ionic strength and has
greater intrinsic activity at room temperature than at 37 degrees C. |
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ISSN: | 0021-9258 1083-351X |
DOI: | 10.1016/s0021-9258(19)49762-7 |