7PHQ
Structure of homo-dimeric Staphylococcus capitis divalent metal ion transporter (DMT) by NabFab-fiducial assisted cryo-EM
Summary for 7PHQ
| Entry DOI | 10.2210/pdb7phq/pdb |
| EMDB information | 13426 |
| Descriptor | NabFab HC, NabFab LC, Divalent metal cation transporter MntH, ... (6 entities in total) |
| Functional Keywords | nabfab, anti-nanobody fab, fiducial, dmt, membrane protein |
| Biological source | synthetic construct More |
| Total number of polymer chains | 10 |
| Total formula weight | 247482.46 |
| Authors | Bloch, J.S.,Mukherjee, S.,Kowal, J.,Kossiakoff, A.A.,Locher, K.P. (deposition date: 2021-08-18, release date: 2021-09-01, Last modification date: 2024-11-20) |
| Primary citation | Bloch, J.S.,Mukherjee, S.,Kowal, J.,Filippova, E.V.,Niederer, M.,Pardon, E.,Steyaert, J.,Kossiakoff, A.A.,Locher, K.P. Development of a universal nanobody-binding Fab module for fiducial-assisted cryo-EM studies of membrane proteins. Proc.Natl.Acad.Sci.USA, 118:-, 2021 Cited by PubMed Abstract: With conformation-specific nanobodies being used for a wide range of structural, biochemical, and cell biological applications, there is a demand for antigen-binding fragments (Fabs) that specifically and tightly bind these nanobodies without disturbing the nanobody-target protein interaction. Here, we describe the development of a synthetic Fab (termed NabFab) that binds the scaffold of an alpaca-derived nanobody with picomolar affinity. We demonstrate that upon complementary-determining region grafting onto this parent nanobody scaffold, nanobodies recognizing diverse target proteins and derived from llama or camel can cross-react with NabFab without loss of affinity. Using NabFab as a fiducial and size enhancer (50 kDa), we determined the high-resolution cryogenic electron microscopy (cryo-EM) structures of nanobody-bound VcNorM and ScaDMT, both small membrane proteins of ∼50 kDa. Using an additional anti-Fab nanobody further facilitated reliable initial three-dimensional structure determination from small cryo-EM test datasets. Given that NabFab is of synthetic origin, is humanized, and can be conveniently expressed in in large amounts, it may be useful not only for structural biology but also for biomedical applications. PubMed: 34782475DOI: 10.1073/pnas.2115435118 PDB entries with the same primary citation |
| Experimental method | ELECTRON MICROSCOPY (8.45 Å) |
Structure validation
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