Structure-function analysis of the 7B2 CT peptide.
Prohormone convertases play important roles in the proteolytic conversion of many protein precursors. The neuroendocrine protein 7B2 and its 31-residue carboxyl-terminal (CT) peptide potently and specifically inhibit prohormone convertase 2 (PC2). We have analyzed the residues contributing to inhibition using N-terminal truncation and alanine scanning. Removal of more than 3 residues from the amino-terminal end of CT1-18 resulted in a more than 190-fold drop in inhibitory activity, showing that most of the residues between 3 and 18 are required for inhibition. In agreement, an Ala scan indicated that only 4 residues could be replaced with Ala without losing mid-nanomolar inhibitory potency; in particular, Gln7, Gln9, and Asp12 could be Ala-substituted to yield peptides with a similar inhibitory potency to the starting peptide. The all-d-retro-inverso, all-l-inverso, and all-d analogues of CT peptide were completely inactive, indicating that amino acid side chains and the CT peptide main chain interact with PC2. CT peptide inhibition could not be competitively blocked by preincubation with truncated CT peptide forms, supporting an absolute requirement for the Lys-Lys pair in initial binding of the CT peptide to the active site.
Apletalina
E V
EV
Department of Biochemistry, Louisiana State University Health Sciences Center, New Orleans, Louisiana, 70112, USA.
Juliano
M A
MA
Juliano
L
L
Lindberg
I
I
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D051379:Mice; D018832:Molecular Chaperones / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism*; D051859:Neuroendocrine Secretory Protein 7B2; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D010907:Pituitary Hormones / Q000737:chemistry* / Q000378:metabolism*; D045706:Proprotein Convertase 2; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D017384:Sequence Deletion; D013381:Subtilisins / Q000037:antagonists & inhibitors; D014162:Transfection
D018832:Molecular Chaperones; D009419:Nerve Tissue Proteins; D051859:Neuroendocrine Secretory Protein 7B2; D010446:Peptide Fragments; D010907:Pituitary Hormones; D011994:Recombinant Proteins; D013381:Subtilisins; D045706:Proprotein Convertase 2