Sequence requirements for proteolytic cleavage of precursors with paired basic amino acids.
When expressed in COS cells, human prorenin was secreted into the medium without being processed to an active renin. Co-expression of furin, a mammalian homologue of the yeast KEX2 gene product, did not affect proteolytic processing of prorenin. A mutant proreninR-4 constructed by site-directed mutagenesis of Pro (-4) to Arg was not cleaved by an endoprotease in the COS cell. However, proreninR-4 was detectably cleaved to yield the active renin upon co-transfection with furin DNA, indicating that Arg at position -4 is important for recognition and processing by furin in addition to the absolute requirement for paired basic amino acids. Another mutant precursor in which Leu (+1) of proreninR-4 was replaced with Ser was found to be much more efficiently processed than proreninR-4, regardless of co-expression of furin. The results suggest that not only a basic amino acid at position -4 but also Leu at position +1 significantly affect the processing of prorenin catalyzed by the COS cell endoprotease or furin.
Department of Biochemistry, Fukuoka University School of Medicine, Japan.
K
K
Oda
M
M
Ikeda
E
E
Tsuji
M
M
Sohda
N
N
Takami
Y
Y
Misumi
Y
Y
Ikehara
D000595:Amino Acid Sequence; D000599:Amino Acids, Diamino; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004792:Enzyme Precursors / Q000235:genetics* / Q000378:metabolism; D045683:Furin; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D011499:Protein Processing, Post-Translational; D012083:Renin / Q000235:genetics* / Q000378:metabolism; D013381:Subtilisins / Q000235:genetics; D014162:Transfection
D000599:Amino Acids, Diamino; D004792:Enzyme Precursors; D009838:Oligodeoxyribonucleotides; D013381:Subtilisins; D045683:Furin; D012083:Renin