Open AccessDevelopment of a LytE‐based high‐density surface display system in Bacillus subtilis
Author(s) -
Chen ChyiLiang,
Wu SauChing,
Tjia Wai Mui,
Wang Christopher L. C.,
Lohka Manfred J.,
Wong SuiLam
Publication year - 2008
Publication title -
microbial biotechnology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.287
H-Index - 74
ISSN - 1751-7915
DOI - 10.1111/j.1751-7915.2007.00017.x
Subject(s) - bacillus subtilis , cell , cell wall , fusion protein , biology , fusion , biophysics , cell fusion , microbiology and biotechnology , biochemistry , recombinant dna , gene , bacteria , genetics , linguistics , philosophy
Summary The three N‐terminal, tandemly arranged LysM motifs from a Bacillus subtilis cell wall hydrolase, LytE, formed a cell wall‐binding module. This module, designated CWBM LytE , was demonstrated to have tight cell wall‐binding capability and could recognize two classes of cell wall binding sites with fivefold difference in affinity. The lower‐affinity sites were approximately three times more abundant. Fusion proteins with β‐lactamase attached to either the N‐ or C‐terminal end of CWBM LytE showed lower cell wall‐binding affinity. The number of the wall‐bound fusion proteins was less than that of CWBM LytE . These effects were less dramatic with CWBM LytE at the N‐terminal end of the fusion. Both CWBM LytE and β‐lactamase were essentially functional whether they were at the N‐ or C‐terminal end of the fusion. In the optimal case, 1.2 × 10 7 molecules could be displayed per cell. As cells overproducing CWBM LytE and its fusions formed filamentous cells (with an average of nine individual cells per filamentous cell), 1.1 × 10 8 β‐lactamase molecules could be displayed per filamentous cell. Overproduced CWBM LytE and its fusions were distributed on the entire cell surface. Surface exposure and accessibility of these proteins were confirmed by immunofluorescence microscopy.