摘要：Pedobacter heparinus heparin lyase II ( Ph HepII) is composed of N-terminal, central, and C-terminal domains. A long surface loop, designated loop-A, is in the N-terminal domain and is composed of amino acids 84-89. In this study, we deleted two, three, or four residues in loop-A to create Δ86-87, Δ85-87, and Δ84-87 Ph HepII deletion mutants. We hypothesized that the deletions would increase Ph HepII thermostability. After heating purified Ph HepII enzymes at 45 °C for 5 min, 6.1 % of the enzyme activity remained in wild-type Ph HepII, whereas 10.6 % of the enzyme activity remained in Δ86-87 Ph HepII. The results indicated that the deletion caused a significant decrease in the activity, although Δ86-87 Ph HepII is slightly more thermostable than wild-type Ph HepII. In addtion, Δ85-87 and Δ84-87 Ph HepII had weak or no enzyme activity, even when unheated. Circular dichroism spectra showed that Δ85-87 Ph HepII was properly folded. These results suggest that the flexibility of loop-A is important for Ph HepII enzyme activity.
