Table 1.
Labels and amino acid sequences of membrane scaffold proteins used for self-assembly of Nanodiscs (adopted with permission from (15)).
| Abbreviation | Description | Amino Acid Sequence |
|---|---|---|
| H1 | Helix 1 | LKLLDNWDSVTSTFSKLREQLG |
| H1 (1-11) | Truncated Helix 1 | STFSKLREQLG |
| H1 (l-17) | Truncated Helix 1 | REQLG |
| H2 | Helix 2 | PVTQEFWDNLEKETEGLRQEMS |
| H3 | Helix 3 | KDLEEVKAKVQ |
| H4 | Helix 4 | PYLDDFQKKWQEEMELYRQKVE |
| H5 | Helix 5 | PLRAELQEGARQKLHELQEKLS |
| H6 | Helix 6 | PLGEEMRDRARAHVDALRTHLA |
| H7 | Helix 7 | PYSDELRQRLAARLEALKENGG |
| H8 | Helix 8 | ARLAEYHAKATEHLSTLSEKAK |
| H9 | Helix 9 | PALEDLRQGLL |
| H10 | Helix 10 | PVLESFKVSFLSALEEYTKKLNTQ |
| FX | Original N-terminus | MGHHHHHHIEGR |
| TEV | Modified N-terminus | MGHHHHHHH DYDIPTTENLYFQG |
| Scaffold variant |
protein | Abbreviated namea |
Composition |
|---|---|---|---|
| 1 | MSP1 | FX-H1-H2-H3-H4-H5-H6-H7-H8-H9-H10 | |
| 2 | MSP2 | FX-H1- H2-H3-H4-H5-H6-H7-H8-H9-H10-GT- H1-H2-H3-H4-H5-H6-H7-H8-H9-H10 | |
| 3 | MSP1E1 | FX-H1-H2-H3-H4-H4-H5-H6-H7-H8-H9-H10 | |
| 4 | MSP1E2 | FX-H1-H2-H3-H4-H5-H4-H5-H6-H7-H8-H9-H10 | |
| 5 | MSP1E3 | FX-H1-H2-H3-H4-H5-H6-H4-H5-H6-H7-H8-H9-H10 | |
| 6 | MSP1D1 | TEV- H1 (1-11)- H2-H3-H4-H5-H6-H7-H8-H9-H10 | |
| 7 | MSP1D2 | TEV- H2-H3-H4-H5-H6-H7-H8-H9-H10 | |
| 8 | MSP2N1 | TEV- H1 (l-11)- H2-H3-H4-H5-H6-H7-H8-H9-H10-GT- H1 (1-11)- H2-H3-H4-H5-H6-H7-H8-H9-H10 | |
| 9 | MSP2N2 | TEV- H1 (1-11)- H2-H3-H4-H5-H6-H7-H8-H9-H10-GT- H2-H3-H4-H5-H6-H7-H8-H9-H10 | |
| 10 | MSP2N3 | TEV- H1 (1-11)- H2-H3-H4-H5-H6-H7-H8-H9-H10-GT- H1 (1-17)- H2-H3-H4-H5-H6-H7-H8-H9-H10 |
Design of the membrane scaffold proteins.
Abbreviated names: (1) MSP1, membrane scaffold protein 1 with N-terminal hexahistidine tag, followed by Factor X recognition site.
(2) MSP2, fusion of two MSP1 molecules with short Glycine-Threonine linker and N-terminal hexahistidine tag, followed by Factor X recognition site.
(3–5) MSP1E1, MSP1E2, MSP1E3, extended membrane scaffold proteins, obtained via insertion of one (residues 56-77 of original MSP1), two (residues 56-99), or three (residues 56-121) extra 22-mer amphipathic helices after the residue Q55 into original MSP1 (1) sequence, as described (10). All have N-terminal hexahistidine tag and Factor X recognition site.
(6) MSP1D1, MSP1 with the first 11 N-terminal amino acids removed which contains the N-terminal heptahistidine tag and TEV protease recognition site with linker.
(7) MSP1D2, MSP1 with the first 22 N-terminal amino acids removed contains the N-terminal heptahistidine tag and TEV protease recognition site with linker.
(8–10) New MSP2 proteins based on MSP1 deletion mutants rather than original MSP1. All three have the same first MSP1D1 molecule, but vary in extent of truncation in the second MSP.