Abstract
BackgroundThe Drosophila melanogaster Serpin 42 Da gene (previously Serpin 4) encodes a serine protease inhibitor that is capable of remarkable functional diversity through the alternative splicing of four different reactive centre loop exons. Eight protein isoforms of Serpin 42 Da have been identified to date, targeting the protease inhibitor to both different proteases and cellular locations. Biochemical and genetic studies suggest that Serpin 42 Da inhibits target proteases through the classical serpin ‘suicide’ inhibition mechanism, however the crystal structure of a representative Serpin 42 Da isoform remains to be determined.ResultsWe report two high-resolution crystal structures of Serpin 42 Da representing the A/B isoforms in the cleaved conformation, belonging to two different space-groups and diffracting to 1.7 Å and 1.8 Å. Structural analysis reveals the archetypal serpin fold, with the major elements of secondary structure displaying significant homology to the vertebrate serpin, neuroserpin. Key residues known to have central roles in the serpin inhibitory mechanism are conserved in both the hinge and shutter regions of Serpin 42 Da. Furthermore, these structures identify important conserved interactions that appear to be of crucial importance in allowing the Serpin 42 Da fold to act as a versatile template for multiple reactive centre loops that have different sequences and protease specificities.ConclusionsIn combination with previous biochemical and genetic studies, these structures confirm for the first time that the Serpin 42 Da isoforms are typical inhibitory serpin family members with the conserved serpin fold and inhibitory mechanism. Additionally, these data reveal the remarkable structural plasticity of serpins, whereby the basic fold is harnessed as a template for inhibition of a large spectrum of proteases by reactive centre loop exon ‘switching’. This is the first structure of a Drosophila serpin reported to date, and will provide a platform for future mutational studies in Drosophila to ascertain the functional role of each of the Serpin 42 Da isoforms.
Highlights
The Drosophila melanogaster Serpin 42 Da gene encodes a serine protease inhibitor that is capable of remarkable functional diversity through the alternative splicing of four different reactive centre loop exons
Specific recognition of the reactive centre loop (RCL) by the target protease is primarily defined by the sequence of the RCL from the P15 to P3’ positions, albeit studies have shown a role for other exosites in determining protease-inhibitor recognition [1,2,3]
The structures are of high quality; the Serpin 42 Da (Spn42Da)-1 structure refined to a Rwork/Rfree of 16.51% and 18.81% respectively, and the Spn42Da-2 structure to a Rwork/ Rfree of 16.58% and 18.54% respectively
Summary
The Drosophila melanogaster Serpin 42 Da gene (previously Serpin 4) encodes a serine protease inhibitor that is capable of remarkable functional diversity through the alternative splicing of four different reactive centre loop exons. Eight protein isoforms of Serpin 42 Da have been identified to date, targeting the protease inhibitor to both different proteases and cellular locations. Within Drosophila there are over 20 inhibitory serpins, Serpin structures are typified by a meta-stable native state, with a solvent exposed RCL that serves as ‘bait’ to bind and inhibit the target protease [1]. Within the final inhibitory complex, the serpin is in a hyper-stable conformation with the ‘hinge’ region of RCL forming the top of the central 4th strand of β-sheet A. This conformation can spontaneously occur upon cleavage of the RCL loop, forming the stable ‘cleaved’ serpin conformation [1]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
