A new member of family 8 polysaccharide lyase chondroitin AC lyase (PsPL8A) from Pedobacter saltans displays endo- and exo-lytic catalysis

Chondroitin lyases are therapeutically important enzymes. The functional aspects of a chondroitin AC lyase (PsPL8A) from Pedobacter saltans DSM12145 were investigated. PsPL8A was cloned in to pET28a(+) vector, expressed in E. coli BL-21(DE3) cells exhibited a molecular size of approximately, 77 kDa. PsPL8A displayed maximum activity with chondroitin 4-sulphate, C4S (489 U mg⁻¹) followed by chondroitin 6-sulphate, C6S (214 U mg⁻¹) and hyaluronic acid (43.2 U mg⁻¹). PsPL8A was maximally active at 39 °C and pH 7.2. 100 mM Na⁺ and 20 mM Ca²⁺ ions enhanced the activity of PsPL8A by 2-fold. The time dependent TLC analysis of PsPL8A degraded products of C4S revealed the presence of higher degree of polymerization (DP) chondroitin sulphate (CS) oligosaccharides at initial stage, but after 1 h, only ΔC4S disaccharide was produced as the major product. This result displayed that PsPL8A follows initially a concomitant endo- and exo-lytic mode which finally shifted to exolytic mode of catalysis. The oligosaccharides released were identified as di-, hexa-, octa- and dodeca-saccharide by ESI–MS analysis. The ΔC4S disaccharide showed a peak at m/z 458 (ESI–MS) while in MS/MS mode it gave the peak at m/z 300. ESI–MS/MS, ¹H- and ¹³C- NMR analyses confirmed the structure of ΔC4S disaccharide product obtained after 24 h reaction of C4S with PsPL8A. The enzyme reported in present study can be used for cancer mitigation, spinal cord injury treatment and CS oligosaccharides production which act as anti-inflammatory agents. This is the first study reporting the cloning and expression of chondroitin AC lyase from Pedobacter saltans DSM 12145.