|Title||Linker editing of pneumococcal lysin ClyJ conveys improved bactericidal activity.|
|Publication Type||Journal Article|
|Year of Publication||2019|
|Authors||Yang, H, Luo, D, Etobayeva, I, Li, X, Gong, Y, Wang, S, Li, Q, Xu, P, Yin, W, He, J, Nelson, DC, Wei, H|
|Journal||Antimicrob Agents Chemother|
|Date Published||2019 Nov 25|
is a leading human pathogen uniquely characterized by choline moieties on the bacterial surface. Our previous work reported a pneumococcal-specific chimeric lysin, ClyJ, which combines the CHAP (cysteine, histidine-dependent amidohydrolase/peptidase) enzymatically active domain (EAD) from the PlyC lysin and the cell-wall binding domain (CBD) from the phage SPSL1 lysin, which imparts choline binding specificity. Herein, we demonstrate that the lytic activity of ClyJ can be further improved by editing the linker sequence adjoining the EAD and CBD domains. Keeping the net charge of the linker constant, we constructed three ClyJ variants containing different lengths of linker sequence. Circular dichroism showed that linker editing only poses minor effects on the folding of EAD and CBD domains. However, thermodynamic examination combined with biochemical analysis demonstrated that one variant, ClyJ-3, with the shortest linker, displayed improved thermal stability and bactericidal activity, as well as reduced cytotoxicity. In a pneumococcal mouse infection model, ClyJ-3 showed significant protective efficacy compared to the ClyJ parental lysin or the Cpl-1 lysin, with 100% survival at a single ClyJ-3 intraperitoneal dose of 100 μg/mouse. Moreover, a ClyJ-3 dose of 2 μg/mouse had the same efficacy as a ClyJ dose of 40 μg/mouse, suggesting a 20-fold improvement Taken together, the present study not only conveys a promising pneumococcal lysin with improved potency, i.e. ClyJ-3, but also implies that, for the first time, the linker sequence plays an important role in determining the activity of a chimeric lysin, providing insight for future lysin engineering studies.
|Alternate Journal||Antimicrob. Agents Chemother.|