Abstract
A glycosyl hydrolase produced by Pseudomonas aeruginosa, PslG, has become a promising candidate for biofilm treatment because of its ability to inhibit and disperse biofilms by disrupting exopolysaccharide matrix at nanomolar concentrations. However, as a protein, PslG used for treatment may be degraded by the ubiquitous proteases (of which trypsin-like serine proteases are a major group) secreted by human cells. This would lead to an insufficient effective concentration of PslG. Here, based on the result of liquid chromatography–tandem mass spectrometry (LC-MS/MS) and structural analysis, we generate a PslG mutant (K286A/K433S) with greatly enhanced trypsin resistance. This measure raises IC50 (the concentration of trypsin that can degrade 50% of protein in 30 min at 37°C) from 0.028 mg mL–1 of the wild-type PslG to 0.283 mg mL–1 of PslGK286A/K433S. In addition, biofilm inhibition assay shows that PslGK286A/K433S is much more efficient than wild-type PslG in the presence of trypsin. This indicates that PslGK286A/K433S is a better biofilm inhibitor than wild-type PslG in clinical use where trypsin-like proteases widely exist.
Highlights
Biofilms are highly structured matrix-enclosed bacterial communities adherent to surfaces (Costerton et al, 1995; Stoodley et al, 2002)
We found that the double-residue mutant PslGK 286A/K 433S selected from 35 mutants showed greatly increased resistance to one of the most widespread proteases— trypsin, compared with the wild-type PslG
Through a combined experimental and computational approach, we have developed a trypsinresistant variant of PslG, which is a promising candidate for clinical and environmental biofilm treatment, the immune response to PslG in clinical usage still needs more research
Summary
Biofilms are highly structured matrix-enclosed bacterial communities adherent to surfaces (Costerton et al, 1995; Stoodley et al, 2002). Biofilm bacteria have extremely enhanced resistance to antibiotic treatments and host immune responses compared with their planktonic living states (Stewart and Costerton, 2001; Stewart and Franklin, 2008). P. aeruginosa can cause both acute and chronic infections, which could be life-threatening. Chronic infection is more difficult to eradicate because of the formation of biofilms and elevated antibiotic resistance. P. aeruginosa can colonize on many medical implants such as catheters, ventilator tubes, and contact lenses, which is the major cause of hospital infections (Ahiwale et al, 2011). P. aeruginosa is the leading cause of chronic respiratory infection and lung infections in patients with cystic fibrosis (Waters and Goldberg, 2019). Amounting to 60–70% contact lens-related keratitis is caused by P. aeruginosa, which causes global blindness and visual impairment (Choy et al, 2008)
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