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-cyclodextrin glucanotransferase, while secretion efficiency was higher than USP45, total yield -cyclodextrin glucanotransferase, although secretion efficiency was higher than USP45, total yield was found to become reduced [26], as a result demonstrating the complex effects brought upon by SPs, not merely around the secretion of heterologous proteins, but on total protein yield at the same time. Apart from SPs, past literature have reported quite a few other techniques which have verified to enhance SE in L. lactis such as the usage of synthetic LEISSTCDA propeptide sequence (SPs are followed by a propeptide sequence which can be cleaved right after translocation to make the mature protein) [23], and the use of a hrtA mutant strains (the only reported cell surface proteolytic housekeeping gene) [22]. In an additional technique, it was shown that the secretion yield of some heterologous proteins is usually improved in L. lactis when co-expressed with B. subtilis PrsA protein, that is a surface anchored protein with chaperon-like functions and happen to be shown to decrease degradation of exported proteins [27].Surface display systemsThe thick and rigid cell wall of gram-positive bacteria too as the lack of an outer membrane envelope has produced them suitable for the cell surface show of proteins. Displaying proteins on bacterial cell wall makes it possible for the bacteria to act as carriers of proteins, particularly antigens, and let interaction of displayed proteins with targeted environments. There are 5 distinct types of protein anchors described in lactic acid bacteria; (1) transmembrane anchors: (2) lipoprotein anchors which binds to the cell membrane; (3) LPXTG-type cell wall anchoring domains; (4) AcmA-repeats anchor domain; (5) S-layer protein attachments that are bound to cell wall elements [28]. In L. lactis, probably the most typically employed strategy for surface display of proteins is by way of the LPXTG sortingsignal of surface-associated proteins that are recognized by the sortase enzyme, and covalently bound towards the cell wall. In this technique, the anchoring mechanism relies around the sortase activity as this membrane-anchored enzyme cleaves the sorting signal with the target protein at its pentapeptide motif (LPXTG) and promotes covalent anchoring on the target protein to the cell wall [29, 30]. However, non-covalent binding of cell surface proteins using lysin motifs (LysM) are also alternatively used, with the LysM of your autolysin AcmA being the most widespread [31]. More interestingly, non-covalent binding of antigens/proteins using AcmA has been shown to permit trans surface display, where proteins are displayed from the outside of L. lactis host cells, as we have previously shown [32]. Employing this strategy, expression of heterologous proteins may be performed in a non-lactococcal host (e.g. E. coli), purified and bound non-covalently towards the lactococcal cell wall simply by mixing the purified heterologous proteins to lactococcal cell cultures. More importantly, this enables the lactococcal cells to carry heterologous proteins with out becoming genetically modified, a technique which have also been demonstrated with Newcastle disease virus hemagglutinin-neuraminidase (HN) protein for precise targeting of breast cancer cells [33]. Also, eukaryotic proteins which need post-translational modifications may also be expressed in eukaryotic hosts, and subsequently attached to L. lactis for delivery [34]. A variation of this process utilizes GEM (gram-positive enhancer matrix) particles that are killed non-recombinant lactococcal cells devoid of most intact cell wall components and intracellular components.