Peptidase leader

This enzyme [EC 3.4.21.89], also known as signal peptidase I and phage-procoat-leader peptidase, catalyzes the hydrolysis of N-terminal leader sequences from secreted and periplasmic protein precursors. It acts on a single bond Ala-Ala in the m-13 phage procoat protein and creates the signal (leader) peptide and coat protein. It is a member of the peptidase family S26 but is unaffected by inhibitors of most serine peptidases. 

This enzyme [EC 3.4.99.35] (better known as signal peptidase II and also known as bacterial leader peptidase I) catalyzes the cleavage of N-terminal leader sequences from membrane prolipoproteins. 

Bacterial cell wall synthesis inhibitor, d-ALANINE-d-ALANINE LIGASE BACTERIAL LEADER PEPTIDASE I Bacterial reorientation,... 

B) Cleavage of a polypeptide loop formed as in (A) by a leader peptidase to give a polypeptide chain anchored by a positively charged cluster near its C terminus. (C) Membrane topology of the E. coli leader peptidase. The active site is in the periplasmic domain. See Tschantz et al.5S0... 

RE Dalbey, W Wickner. Leader peptidase catalyzes the release of exported proteins from the outer surface of the Escherichia coli plasma membrane. J Biol Chem 260 15925-15931, 1985. 

T Inada, DL Court, K Ito, Y Nakamura. Conditionally lethal amber mutations in the leader peptidase gene of Escherichia coli. J Bacteriol 171 585-587, 1989. 

G. Schneider and P. Wrede, Biophys. /., 66,335 (1994). The Rational Design of Amino Acid Sequences by Artificial Neural Networks and Simulated Molecular Evolution De Novo Design of an Idealized Leader Peptidase Cleavage Site. 

FIGURE 2. Processing of lumenal proteins by TPP and E.coli leader peptidase. Wheat pre-33 K and pre-23 K (lanes 1) were incubated with pea TPP (lanes 2) or leader peptidase (lanes 3). 

On-Bead Libraries for the Discovery of Substrates for Proteases Using Pure Enzyme Escherichia coli Leader Peptidase... 

Fig. 1. A substrate for E. coli leader peptidase is shown, 1, with a quenching Dabsyl group and a fluorescent lucifer yellow attached suitable for FRET-based assay. This substrate was a template for design of a peptide library as shown in structure 2. The arrow indicates the position of cleavage for the peptidase between Ala-Ala/Pro. X represents randomized positions in which any of 19 different natural amino acids are coupled (all except Cys). Note that Dabsyl is on the epsilon amino group of the N-terminal lysyl residue, K.

Even though library 2 (Fig. 1) was designed for E. coli leader peptidase, it was found to be a useful library for screening against other proteases. In the example given, it was used to screen for napsin A substrates, a mammalian protease expressed in kidney and lung (8,9). The protease inhibitors E64 and... 

In the case of E. coli leader peptidase, 45 brightly fluorescent beads were isolated and analyzed by Edman chemistry sequencing (Fig. 3). 

The expected sequence of AXAA was obtained. Of 45 beads, 39 were cleaved between the Ala-Ala. The remaining 6 beads were cleaved between X3-Ala, in which case Ala was found in the Xj position. Thus, essentially all of the isolated beads were satisfactory substrates for the leader peptidase. There was modest enrichment of certain amino acids in some of the positions, as expected. Resynthesis of 7 of the 39 sequences selected at random yielded surrogate substrates with a range of relative catalytic efficiencies from 0.3- to 19-fold compared to the known substrate 1 (Fig. 1). Two of the substrates were 11- and 19-fold better (8). 

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