Characterization of Dipeptidyl Peptidase IV Enzyme Activity in Serum of the Komodo Dragon (Varanus komodoensis)

Aims: Soluble serum dipeptidyl peptidase IV (DPPIV) is a protease that cleaves dipeptides from proteins that have alanine or proline next to the N-terminal amino acid. This enzyme demonstrates substantial immune function by regulating T-lymphocyte activity, T-cell chemotaxis, growth, and proliferation during an inflammatory response. The goal of this study was to characterize DPPIV activity in the serum of the Komodo dragon (Varanus komodoensis).

Study Design: Serum from captive Komodo dragons were pooled and the DPPIV activities were determined at a range of serum dilutions, temperatures, and time points. The effects of Diprotin A, a specific DPPIV enzyme inhibitor, on the generation of fluorescent product were also determined. All samples were analyzed in quadruplicate such that meaningful statistical evaluations could be conducted.

Place and Duration of Study: Serum was collected from eight Komodo dragons at the San Antonio Zoo (n=5) and Houston Zoo (n=3) in June of 2012. The samples were analyzed for DPPIV enzyme activity in the Department of Chemistry at McNeese State University in Lake Charles, Louisiana, USA

Methodology: We used Ala-Pro-AFC, a dipeptide conjugated to a fluorescent probe via an amide linkage, to measure the activity of DPPIV in the serum of Komodo dragons (Varanus komodoensis). The fluorescent intensity of the product formed was measured at excitation and emission wavelengths of 395 and 530 nm, respectively, in a fluorimeter.

Results: Incubation of different volumes of serum from the Komodo dragon with Ala-Pro-AFC resulted in a volume-dependent increase in fluorescent intensity, which was decreased in a concentration-dependent manner by diprotin A, a specific inhibitor of DPPIV activity. Kinetic analysis showed that the DPPIV enzyme activity was detectable after five minutes, and that was nearly linear for three hours. A thermal profile showed that Komodo dragon DPPIV exhibited dramatically reduced activities at low temperatures (5-10°C), but activity increased linearly with temperature and was maximal at the highest temperature tested (40°C).

Conclusion: These results from this study indicate that Komodo dragons exhibit considerably high serum DPPIV activities, which are likely to contribute to T-cell activation and function, and act as a bridge between innate and adaptive immunity in these ancient vertebrates.