Human Single-Chain Antibodies That Neutralize Elastolytic Activity of <i>Pseudomonas aeruginosa</i> LasB

• Main Authors: Sirijan Santajit, Thida Kong-ngoen, Manas Chongsa-Nguan, Usa Boonyuen, Pornpan Pumirat, Nitat Sookrung, Wanpen Chaicumpa, Nitaya Indrawattana
• Format: Article
• Language: English
• Published: MDPI AG 2021-06-01
• Series: Pathogens
• Subjects: elastase; elastolytic activity; human single-chain antibody; HuscFv; <i>Pseudomonas aeruginosa</i>; LasB
• Online Access: https://www.mdpi.com/2076-0817/10/6/765

LasB (elastase/pseudolysin) is an injurious zinc-metalloprotease secreted by the infecting <i>Pseudomonas aeruginosa</i>. LasB is recognized as the bacterial key virulence factor for establishment of successful infection, acquisition of nutrients, dissemination, tissue invasion, and immune modulation and evasion. LasB digests a variety of the host tissue proteins, extracellular matrices, as well as components of both innate and adaptive immune systems, including immunoglobulins, complement proteins, and cytokines. Thus, this enzyme is an attractive target for disarming the <i>P. aeruginosa.</i> This study generated human single-chain antibodies (HuscFvs) that can neutralize the elastolytic activity of native LasB by using phage display technology. Gene sequences coding HuscFvs (<i>huscfvs</i>) isolated from HuscFv-displaying phage clones that bound to enzymatically active LasB were sub-cloned to expression plasmids for large scale production of the recombinant HuscFvs by the <i>huscfv</i>-plasmid transformed <i>Escherichia coli</i>. HuscFvs of two transformed <i>E. coli</i> clones, i.e., HuscFv-N42 and HuscFv-N45, neutralized the LasB elastolytic activities <i>in vitro</i>. Computer simulation by homology modeling and molecular docking demonstrated that antibodies presumptively formed contact interfaces with the LasB residues critical for the catalytic activity. Although the LasB neutralizing mechanisms await elucidation by laboratory experiments, the HuscFvs should be tested further towards the clinical application as a novel adjunctive therapeutics to mitigate severity of the diseases caused by <i>P. aeruginosa</i>.