Proteolytic system text description

PROTEOLYTIC SYSTEM

The proteolytic system of lactic acid bacteria often consists of an extracellular, peptidoglycan-bound proteinase which cleaves milk proteins into oligo-, tri- and dipeptides (11). These peptides can then enter the cell via peptide transport systems and are further degraded into amino acids by dedicated peptidases. However, in many LAB species or strains there is no extracellular proteinase (11).

 Extracellular proteases

L. plantarum was not previously known to encode an extracellular serine proteinase of the subtilisin-like family (also called subtilase), as found in several other lactic acid bacteria, either on their chromosome or on a plasmid. This cell-envelope bound subtilase allows bacteria to use proteins and oligopeptides as growth substrates. In the present study, 9 L. plantarum strains were found to have an extracellular subtilase (OG_3299), cell-envelope bound by an LPXTG anchor, i.e. in strains NIZO1837, NIZO2260, NIZO2259, NIZO2806, NIZO3400, NIZO3893, NIZO2802 (pseudogene), P8 and 16 (pseudogene). All of these appear to be plasmid-encoded.

There are several other known families of extracellular proteases and peptidases in lactic acid bacteria (see Secretome section). L. plantarum WCFS1 encodes 4 different extracellular zinc metalloproteinases (OG_2266, OG_3046, OG_2488, OG_2279), all of the M10 family. The first two are present in all strains (a few as pseudogenes), one zinc metalloproteinase (OG_2488) is missing in 12 strains and one zinc metalloproteinase (OG_2279) is present in all strains, except for NIZO1838 and NIZO2256 strains. Each strain has 6-9 genes encoding membrane-bound proteases of the CAAX family, which are presumably all extracellular. These proteins are also called self-immunity proteins as they are presumed to play a role in defense against their own bacteriocins like plantaricins, although only 2 genes are in the plantaricin gene cluster. One extracellular S41 family carboxy-terminal processing proteinase (OG_1831)  is present in all strains, whereas another (OG_2365) is absent or a pseudogene in 6 strains. No other differences were found in extracellular proteases of the L. plantarum strains. It is possible that some of the extracellular proteins of unknown function are also proteases.

 

Intracellular proteases/peptidases

An intracellular protease/endopeptidase of the subtilase family (OG_3734) is found in 4 strains, i.e. ER, NIZO2830, NIZO2831, and JDM1. Surprisingly, the 4 protein sequences are 100% identical.

Most intracellular peptidases of known specificity (11) are present and complete in all L. plantarum genomes – only a few are putative pseudogenes. Two variants of PepC and PepF are found, and 4 variants of PepD encoding a dipeptidase. Four variants of prolyl peptidases are found, of which two are present in all strains (OG_548, OG_1874) and two only in about half the strains (OG_2929, OG_2906). One of the latter prolyl peptidases (OG_2929) is always encoded adjacent to a transport protein for proline-containing peptides (OG_2853). Absent in all genomes are the peptidases Pcp, PepS, PepA, and PepE/PepG, which are more commonly found in other lactic acid bacteria (11).

There are another 4 putative intracellular zinc-dependent metallopeptidases, generally membrane-bound, present in all strains (OG_1535, OG_554, OG_1722, OG_1779). Their function is unknown.

In addition, there are a number of housekeeping proteases generally involved in regular protein turnover, such as HtrA, Lon, and LexA, and the ATP-dependent Clp and Hsl proteases. These appear to be present and complete in all 54 strains. 

 

Peptide transporters

Oligopeptide ABC transporter

An Opp system (oppABCDF) is present and complete in all strains (orthologs of lp_1261-1265 in strain WCFS1). In addition, all strains have 4-5 intact extra genes encoding OppA paralogs for binding of oligopeptides, possibly with different specificity.

Dipeptide/tripeptide/oligopeptide transporter

An additional complete dipeptide/tripeptide/oligopeptide ABC transporter (OGs 3925, 3763, 3764) is present in 5 strains, i.e. NIZO2259, NIZO2264, NIZO2484, NIZO2485, and ZJ316. It is not clear what size peptides are transported.

A complete di/tripeptides ion-linked permease, MFS superfamily (OG_955) is present in all genomes.  Twenty-two strains appear to have a complete transport protein or proline-containing amino acid or peptides (OG_2853).

 

11.      Liu M, Bayjanov JR, Renckens B, Nauta A. The proteolytic system of lactic acid bacteria revisited: a genomic comparison. BMC …. 2010.