<p>This study aimed to clone and express chitinase genes from <i>Bacillus proteolyticus</i> strains and characterize the enzymatic properties of recombinant enzymes. <i>Bacillus proteolyticus</i> was isolated from the body of <i>Dermacentor nuttalli</i> and renamed IMH/B-1. Chitin-degrading enzymes were screened via clear zone assay and PCR. The chitinase A gene (<i>ChiA</i>) was successfully cloned, and the recombinant plasmid pET28a-rChiA-DP (Dermatestor-derived Protein) was constructed. Recombinant chitinase protein (rChiA-DP) was expressed in <i>Escherichia coli</i> BL21 using IPTG induction and purified by nickel-nitrilotriacetic acid (Ni–NTA) affinity chromatography. Bioinformatic tools were used to predict the rChiA-DP protein sequences, analyse its enzyme family classification, and identify key amino acid residues in its catalytic domain. The enzymatic activity of rChiA-DP, along with its nematode resistance and antifungal effects on <i>Caenorhabditis elegans</i> (<i>C. elegans</i>) and fungi (<i>Aspergillus</i> sp.), was assessed under varied temperatures, pH, metal ions, salt concentrations and substrates. The amino acid sequence of the rChiA-DP contains a chitin-binding domain (CBD) (substrate binding), a fibronectin type III domain (FN3)(structural stability), and a catalytic domain with a typical TIM-barrel molecular structure (catalytic scaffold). SDS-PAGE analysis revealed that the molecular weight of the rChiA-DP was approximately 74.6&#xa0;kDa, which is consistent with the theoretical predictions. The optimal conditions for rChiA-DP enzyme activity were 40 °C and pH 7.0. Enzyme activity was significantly enhanced by 10&#xa0;mM Ba²⁺, Tris, K⁺, and Li⁺. Organic solvents such as methanol, ethanol, isopropanol, and isoamyl alcohol (10% concentration) also increased the activity. Conversely, positive metal ions such as Cu²⁺, Ni²⁺, Fe³⁺, Zn²⁺ and Mn²⁺ as well as SDS, DMSO, Tween 20/80 significantly inhibited the activity of the rChiA-DP. rChiA-DPs demonstrated varying degrees of decomposition activity against substrates such as colloidal chitin, chitin powder, nematode eggs, nematodes, shrimp shells, and tick eggs, with the highest activity observed for colloidal chitin (7.53 ± 0.86 U/mL). However, it exhibited no degradation activity on chitosan and tick surface. Compared with the heat-inactivated control group‌ and the s-buffer group, the rChiA-DP treatment significantly reduced the survival rate of <i>C. elegans</i> by 50.4% vs. heat-inactivated control (<i>P</i> &lt; 0.01), indicating a potential antiparasitic effect. However, it showed no significant antifungal activity against fungi such as <i>Aspergillus niger</i> or <i>Aspergillus flavus,</i> and the diameter of the inhibition zone was not significantly larger than that of the negative control (<i>P</i> &gt; 0.05). This study successfully prepared tick-derived rChiA-DPs and evaluated their enzymatic activity and anti-nematodal activity, providing enzymatic basis for the design of biopesticides targeting insect cuticle.</p>