## 2 Results
### LapA Domain vWFa Is Involved in the Peptide-Dependent Biofilm of P. putida PaW85
Homogeneity groups are shown above the columns by lower-case letters. Identical letters denote nonsignificant differences (\(p\geq 0.05\)) between averages of biofilm. Multifactorial ANOVA was used for statistical analysis, analysed separately by media. Figure 6: SDS-PAA gels and biofilm of _P. putida_PSm-G-based strains and biofilm. (**A**) SDS-PAA gel electrophoresis of crude cell lysates, 40 ug of total protein is loaded to each lane. The media in which cells were grown for preparing crude cell lysates are shown on the left of the gel image. Cells were grown with and without 1 mM IPTG. The strains used in the analysis are shown above and below the figure. PSm\(\Delta\)lapA and PSm\(\Delta\)lapF strains were used as negative controls for LapA and LapF. Arrows indicate LapA and LapF on the right side of the figure. The inclined arrow indicates an unknown protein complex. (**B**) Biofilm of PSm-G-based strains in M9-0.2 CAA, (**C**) M9-0.2 CAA+tryptone and (**D**) LB in the presence and absence of 1 mM IPTG. An arithmetical average of at least three biological parallels with 95% confidence intervals are given. Homogeneity groups are shown above the columns by lower-case letters. Identical letters denote nonsignificant differences (\(p\geq 0.05\)) between averages of biofilm. Multifactorial ANOVA was used for statistical analysis, analysed separately by media. Figure 7: Expression of _fs_ in F15-based strains by IPTG, the presence of LapA and LapF in cells and biofilm. (**A**) Expression of _fs_ in _P. putida_ F15 by immunoblotting using monoclonal anti-Fis antibodies. 10 \(\upmu\)g of total protein obtained from _P. putida_ F15 grown in LB, M9-0.2 CAA or M9-0.2 CAA+tryptone media was loaded on a 12% SDS-PA gel. The supplementation of 1 mM IPTG is shown by “+” above the lane. 45 ng of purified Fis (6 His) was used as a positive control. Protein marker band sizes are indicated. (**B**) SDS-PAA gel electrophoresis of crude cell lysates, 40 \(\upmu\)g of total protein is loaded to each lane. The media in which cells were grown for preparing crude cell lysates are shown to the left of the gel image. Cells were grown with and without 1 mM IPTG. The strains used in the analysis are shown above and below the figure. PSmAlapA and PSmAlapF strains were used as negative controls for LapA and LapF. LapA and LapF are indicated by arrows on the right side of the figure. Inclined arrows indicate an unknown protein complex. (**C**) Biofilm of _P. putida_ F15-based strains. An arithmetical average of at least three biological parallels with 95% confidence intervals are given. Homogeneity groups are shown above the columns by lower-case letters. Identical letters denote nonsignificant differences (\(p\geq 0.05\)) between averages of biofilm. Multifactorial ANOVA was used for statistical analysis, analysed separately by media. In a polylysine-containing M9-0.2 CAA medium, IPTG did not affect the biofilm of the P-PANB strain (Figure 4E). Still, the biofilm of vWFa domain deletion strains (Avwf\({}^{-}\)) was decreased compared to the original strains from which they were constructed (Figure 4E). Independent of IPTG, the strain P-PANB-Avwf\({}^{-}\) had 1.5 times less biofilm than the P-PANB strain (\(p<0.0001\), Figure 4E). For PaW85 versus P-Avwf\({}^{-}\), deletion of the vWFa domain reduced biofilm 1.2-fold (\(p=0.063\)) and 1.4-fold (\(p<0.001\)) in the absence and presence of IPTG, respectively (Figure 4E). Our previous study showed that adding water-soluble cellulose to a medium enhances _P. putida_ biofilm [19]. Deleting the vWFa domain did not affect the biofilm in the M9-0.2 CAA+cellulose medium; both vWFa-deletion strains behaved similarly to its origin strain, P-Avwf\({}^{-}\) similarly to PaW85 and P-PANB-Avwf\({}^{-}\) similarly to P-PANB (Figure 4F). However, the addition of IPTG to the medium increased 1.2-times biofilms of P-PANB strains compared to cells grown without IPTG, P-PANB (\(p=0.006\)) and the P-PANB-Avwf\({}^{-}\) strain (\(p<0.0001\); Figure 4F).