Reversed phase HPLC analysis of proenkephalin-related and prodynorphin-related end-products in the brain of a urodele amphibian, Ambystoma tigrinum.
Acid extracts of the brain of a urodele amphibian, Ambystoma tigrinum, were screened with radioimmunoassays specific for enkephalin-related products and dynorphin-related products. Following Sephadex G-50 column chromatography a peak of enkephalin-sized immunoreactive material was detected near the total volume of the column. The enkephalin-sized immunoreactivity was further analyzed by reversed phase HPLC. This analysis detected peaks of authentic Met-enkephalin and Leu-enkephalin. However, the molar ratio of Met-enkephalin to Leu-enkephalin in the brain of this amphibian was approximately 80:1. These observations would suggest that the Leu-enkephalin detected in the brain of Ambystoma may be derived from a source other than the Proenkephalin precursor. Neither Met-enkephalin-RGL or Met-enkephalin-RF were detected by radioimmunoassay in brain extracts from this urodele. However, following digestion with trypsin and carboxypeptidase B, a novel peak of C-terminally extended Met-enkephalin was detected. Two peaks of Prodynorphin-related products were also detected following gel filtration chromatography. These immunoreactive forms were detected using antisera specific for alpha-neo-endorphin and dynorphin B(1-13). No immunoreactive forms with antigenic determinants similar to mammalian dynorphin A(1-17) or dynorphin A(1-8) were detected in this species. Reversed phase HPLC analysis indicated that the major form of urodele alpha-neo-endorphin eluted with the same retention time as synthetic mammalian alpha-neo-endorphin. Urodele dynorphin B(1-13)-related immunoreactivity eluted as a single peak, however, this form did not elute with the same retention time as synthetic mammalian dynorphin B(1-13).(ABSTRACT TRUNCATED AT 250 WORDS)
Dores
R M
RM
University of Denver, Department of Biological Sciences, CO 80208.
McDonald
L K
LK
Purdom
L C
LC
Sei
C A
CA
D000557:Ambystoma / Q000235:genetics*; D000818:Animals; D005075:Biological Evolution; D001921:Brain / Q000378:metabolism*; D001923:Brain Chemistry; D002851:Chromatography, High Pressure Liquid; D004745:Enkephalins / Q000235:genetics*; D005260:Female; D005786:Gene Expression Regulation / Q000502:physiology; D008297:Male; D011498:Protein Precursors / Q000235:genetics*
D004745:Enkephalins; D011498:Protein Precursors; C029992:proenkephalin; C033660:preproenkephalin