Molecular determinants of human prorenin processing.
In humans, active renin is generated by the removal of a 43-amino acid prosegment from the zymogen prorenin. This cleavage event is highly specific, occurring at only one of the seven pairs of basic amino acids in the body of preprorenin. This cleavage site selectivity is also displayed by a number of other proteases in vitro and in mouse pituitary AtT-20 cells transfected with a human preprorenin expression vector, suggesting that specificity of cleavage is directed in part by the primary sequence, the higher order structure, or both of prorenin itself. To test this hypothesis, single amino acid mutations were introduced in the region of human preprorenin surrounding the natural cleavage site, and the resultant recombinant proteins were expressed in cultured Chinese hamster ovary and AtT-20 cells. The results suggest that amino acids in addition to the pair of basic amino acids surrounding the cleavage site affect the ability of both trypsin and the endogenous AtT-20 processing enzyme to cleave prorenin. Notably, although a proline at position -4 is essential for processing of prorenin in AtT-20 cells and is correlated with predicted formation of a beta-turn at this position, site-directed mutations suggest that this structural feature in addition to a pair of basic amino acids is not sufficient to lead to proteolytic activation of prorenin. Displacement of sequences surrounding the cleavage site to a position 10 amino acids toward the amino terminus led to partial processing of a mutated prorenin.(ABSTRACT TRUNCATED AT 250 WORDS)
Chu
W N
WN
Metabolic Research Unit, University of California, San Francisco.
Mercure
C
C
Baxter
J D
JD
Reudelhuber
T L
TL
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells / Q000378:metabolism; D002461:Cell Line, Transformed / Q000378:metabolism; D006224:Cricetinae; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011499:Protein Processing, Post-Translational; D051381:Rats; D012083:Renin / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D004792:Enzyme Precursors; D012083:Renin