Role of Amino Acid Sequences Flanking Dibasic Cleavage Sites in Precursor Proteolytic Processing

Abstract

The amino acid sequences flanking 352 dibasic moieties contained in 83 prohormones and pro-proteins listed in a database were examined. Frequency calculations on the occurrence of given residues at positions P, to P: allowed us to delineate a number of features which might be in part responsible for the in vivo discrimination between cleaved and uncleaved dibasic sites. These include the following: amino acids at these positions were characterized by a large variability in composition and properties; no major contribution of a given precursor subsite to endoprotease specificity was observed; some amino acid residues appeared to occupy preferentially certain precursor subsites (for instance, Met in P, and P,, Asp and Ala in P;, Pro in P,, Gly in P, and P: etc.) whereas some others appeared to be excluded.Most amino acid residues occupying the P: position in these precursor cleavage sites were tolerated. But the P-carbon branched side chain residues (Thr, Val, Leu, Ile) and Pro, Cys, Met and Trp were either totally excluded or poorly represented, suggesting that they might be unfavourable to cleavage. The biological relevance of these observations to the efficacy of dibasic cleavage by model propeptide convertases was in vitro tested using both pro-ocytocin convertase and Kex2 protease action on a series of proocytocin related synthetic substrates reproducing the Pro7+Leul5 sequence of the precursor in which the Ala13 residue (Pi in the LysArg-Ala motif) was replaced by various amino acid residues. A good correlation was obtained on this model system indicating that Pi residue of precursor dibasic processing sites is an important feature and may play the role of anchoring motif to S: convertase subsite.We tentatively propose that the present database, and the corresponding model, may be used for further investigation of dibasic endoproteolytic processing of propeptides and pro-proteins.

Neuropeptidome of the Cephalopod Sepia officinalis: Identification, Tissue Mapping, and Expression Pattern of Neuropeptides and Neurohormones during Egg Laying

“…A bradykinin-like neuropeptide precursor gene is expressed in neuron L5 of Aplysia californica . The related neuropeptides have been renamed neuropeptides KY because the N-terminal amino acid is predicted to be a lysine and the C-terminal amino acid is predicted to be a Yamide. , …”

Mammalian Peptide Hormones: Biosynthesis and Inhibition

“…Are there conformational requirements for prohormone convertase substrates that can be used for inhibitor design? Although there are several preferences for prohormone cleavage sites [83,[234][235][236] that can be visualized well by a sequence logo [237,238] (an example is shown in Figure 13.11), examination of sequences around cleaved sites finally could not reveal any consensus primary sequence [239] that is cleaved in all cases. Thus, the question arises, what is the reason for a prohormone convertase to cleave at some basic sites and to leave a lot of other basic sites intact?…”

Sensitive Identification of Known and Unknown Protease Activities by Unsupervised Linear Motif Deconvolution

“…Unlike other PTMs, proteolytic events are centered around a bond between two residues. Moreover, the amino acids found adjacent to proteolytic cleavage sites are characterized by a large variability in composition and properties . In addition to this, the cooperative relationship of successive positions (P4–P4′) on substrate sequences has been established for specific proteases, , strongly highlighting the need to consider the interaction of adjacent amino acids in the study of protease–substrate cleavage specificity.…”

“…Moreover, the amino acids found adjacent to proteolytic cleavage sites are characterized by a large variability in composition and properties. 8 In addition to this, the cooperative relationship of successive positions (P4− P4′) on substrate sequences has been established for specific proteases, 9,10 strongly highlighting the need to consider the interaction of adjacent amino acids in the study of protease− substrate cleavage specificity. Proteolytic data sets therefore lack the fixed central residue required by these tools, even if the sequence alignment is shifted in order to center the sequences around a residue position rather than the cleavage site.…”