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A rapid assay for prolyl 3-hydroxylase activity

โœ Scribed by Juha Risteli; Karl Tryggvason; Kari I. Kivirikko

Publisher
Elsevier Science
Year
1978
Tongue
English
Weight
451 KB
Volume
84
Category
Article
ISSN
0003-2697

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โœฆ Synopsis

An assay is reported for prolyl 3-hydroxylase activity. The method is based on the release of tritiated water (THO) during 3-hydroxylation of a 2,3-T-~proline-labeled (T = tritium) polypeptide substrate in which all prolyl residues recog nized by prolyl 4-hydroxylase have been converted to 4-hydroxyprolyl residues. The formation of THO was essentially linear with enzyme concentration and time, and the K, for the polypeptide substrate was about 3.4 x lo-* M. A linear correlation was found between THO release and the synthesis of 3-hydroxyproline, the latter being analyzed by amino acid analyzer. The assay is simple, rapid, sensitive, and reproducible, and it is specific even in tissue samples containing a large excess of prolyl 4-hydroxylase activity.

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