Inactivity of Recombinant ELA2B Provides a New Example of Evolutionary Elastase Silencing in Humans

Background: The archetypal mammalian elastase (ELA1) is not expressed in the human pancreas, because evolutionary mutations suppressed transcription of the ELA1 gene. Aims: In this study, we tested the theory that the unique duplication of the ELA2 gene in humans might compensate for the loss of ELA1.Methods: Recombinant ELA2A and ELA2B were expressed in Escherichia coli, and their activity was tested on Glt-Ala-Ala-Pro-Leu-p- nitroanilide, DQ elastin and bovine milk protein.Results: Surprisingly, recombinant ELA2B was completely devoid of proteolytic activity, while ELA2A readily hydro- lyzed all three test substrates. Furthermore, ELA2A formed an SDS-resistant complex with α1-antitrypsin, whereas ELA2B did not bind covalently to the inhibitor. Finally, chimeras and point mutations engineered between ELA2A and ELA2B revealed that multiple evolutionary mutations inactivated ELA2B.Conclusions: The results indicate that ELA2B is not an elastase enzyme and confirm that ELA2A is the major elastase in the human pancreas.