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. 2020 May 27;21(11):3773. doi: 10.3390/ijms21113773

Figure 1.

Figure 1

The expression cassette (A), erm-based and alr-based expression vectors (B,C) of C-terminal cell-wall anchor of chitosanase (CsnA) in L. plantarum. (A) Myc tag sequence was fused to the C-terminus of CsnA for protein detection. All parts of the expression cassette can be easily exchangeable using the indicated restriction enzymes (BglII, SalI, MulI, EcoRI). The signal peptide (SP) derived from Lp_0373 was fused to CsnA via SalI site. The short anchor S (85 residues) consists of a linker of 47 residues between the LP × TG motif, which is the consensus sequence LPQTNE in L. plantarum [30,31] (bold, underlined), and the C-terminus of CsnA-Myc with a valine as the fusing point (V in bold). The long anchor L (194 residues) contains a linker of 156 residues upstream of the LP × TG motif including one Mub B2-like domain of 69 residues (mucus binding domain; in italics) fused to CsnA-Myc with a threonine residue (T in bold). (B) Schematic overview of the plasmid encoding CsnA fused to a LP × TG cell wall anchor using different lengths of Lp_1229 derived anchor and Lp_0373 derived SP for secretion. The plasmid contains an erythromycin antibiotic resistance gene (erm) as selection marker. (C) Schematic overview of the plasmid encoding CsnA fused to a LP × TG cell wall anchor using different lengths of Lp_1229 derived anchor and Lp_0373 derived SP for secretion. The plasmid contains alanine racemase gene (alr) as selection marker. See text for more details.