Structural Modelling of the Lumenal Domain of Human GPAA1, the Metallo-Peptide Synthetase Subunit of the Transamidase Complex, Reveals Zinc-Binding Mode and Two Flaps Surrounding the Active Site

Structural Modelling of the Lumenal Domain of Human GPAA1, the Metallo-Peptide Synthetase Subunit of the Transamidase Complex, Reveals Zinc-Binding Mode and Two Flaps Surrounding the Active Site
Title:
Structural Modelling of the Lumenal Domain of Human GPAA1, the Metallo-Peptide Synthetase Subunit of the Transamidase Complex, Reveals Zinc-Binding Mode and Two Flaps Surrounding the Active Site
Other Titles:
Biology Direct
Publication Date:
29 September 2020
Citation:
Su, C.TT., Sinha, S., Eisenhaber, B. et al. Structural modelling of the lumenal domain of human GPAA1, the metallo-peptide synthetase subunit of the transamidase complex, reveals zinc-binding mode and two flaps surrounding the active site. Biol Direct 15, 14 (2020). https://doi.org/10.1186/s13062-020-00266-3
Abstract:
Background: The transamidase complex is a molecular machine in the endoplasmic reticulum of eukaryotes that attaches a glycosylphosphatidylinositol (GPI) lipid anchor to substrate proteins after cleaving a C-terminal propeptide with a defined sequence signal. Its five subunits are very hydrophobic; thus, solubility, heterologous expression and complex reconstruction are difficult. Therefore, theoretical approaches are currently the main source of insight into details of 3D structure and of the catalytic process. Results: In this work, we generated model 3D structures of the lumenal domain of human GPAA1, the M28-type metallo-peptide-synthetase subunit of the transamidase, including zinc ion and model substrate positions. In comparative molecular dynamics (MD) simulations of M28-type structures and our GPAA1 models, we estimated the metal ion binding energies with evolutionary conserved amino acid residues in the catalytic cleft. We find that canonical zinc binding sites 2 and 3 are strongest binders for Zn1 and, where a second zinc is available, sites 2 and 4 for Zn2. Zinc interaction of site 5 with Zn1 enhances upon substrate binding in structures with only one zinc. Whereas a previously studied glutaminyl cyclase structure, the best known homologue to GPAA1, binds only one zinc ion at the catalytic site, GPAA1 can sterically accommodate two. The M28-type metallopeptidases segregate into two independent branches with regard to one/two zinc ion binding modality in a phylogenetic tree where the GPAA1 family is closer to the joint origin of both groups. For GPAA1 models, MD studies revealed two large loops (flaps) surrounding the active site being involved in an anti-correlated, breathing-like dynamics. Conclusions: In the light of combined sequence-analytic and phylogenetic arguments as well as 3D structural modelling results, GPAA1 is most likely a single zinc ion metallopeptidase. Two large flaps environ the catalytic site restricting access to large substrates.
License type:
http://creativecommons.org/licenses/by/4.0/
Funding Info:
There was no specific funding for research done.
Description:
This is a post-peer-review, pre-copyedit version of an article published in Biology Direct. The final authenticated version is available online at: http://dx.doi.org/10.1186/s13062-020-00266-3
ISSN:
1745-6150
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