Key resources table.
| Reagent type (species) or resource |
Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Gene (human) | Akt1 | Addgene and DOI:10.1093/nar/gkh238 |
Addgene # 9021 | |
| Strain, strain background (E. coli) | Rosetta 2(DE3) pLysS | Novagen | Cat. No.: 71400 | Competent cells |
| Strain, strain background (E. coli) | DH10Bac | Invitrogen | Cat. No.: 18297010 | Competent cells |
| Cell line (insect cell) | Sf21 | Invitrogen | 11497–013 | |
| Cell line (insect cell) | f9 | Invitrogen | 11496–015 | |
| Cell line (human) | Akt1/2-/- HCT116 | DOI:10.1073/pnas.0914018107 | Colon cancer cell line; The Akt1-/-and Akt2-/-HCT116 colon cancer cell line was a gift from Dr. Bert Vogelstein (Johns Hopkins University) (Ericson et al., 2010). These cells were authenticated by western blot showing the absence of Akt and by the lack of signaling response to growth factors. They were also shown to be mycoplasma-free by PCR testing. | |
| Antibody | pan-Akt (11E7) (Rabbit monoclonal) |
Cell Signaling Technology | Cat. No.: 4685S, RRID:AB_10698888 | WB (1:1000 for cell-based assays; 1:20000 for activation assays) |
| Antibody | Akt phospho-Thr308 (D25E6) (Rabbit monoclonal) |
Cell Signaling Technology | Cat. No.: 13038S, RRID:AB_2629447 |
WB (1:1000, for cell-based assays; 1:10000 for activation assays) |
| Antibody | Akt phospho-Ser473 [EP2109Y] (Rabbit monoclonal) |
AbCam | Cat. No.: ab81283, RRID:AB_2224551 |
WB (1:1000) |
| Antibody | GAPDH (14C10) (Rabbit monoclonal) | Cell Signaling Technology | Cat. No.: 2118S, RRID:AB_561053 |
WB (1:5000) |
| Antibody | HRP conjugated, anti-Rabbit IgG (Goat monoclonal) | Cell Signaling Technology | Cat. No.: 7074S, RRID:AB_2099233 | WB (1:5000) |
| Chemical compound, drug | Deuterium oxide (D, 99.8%) | Cambridge Isotope Laboratories | DLM-4–99.8-1000 | |
| Chemical compound, drug | Ammonium chloride (15N, 99%) | Cambridge Isotope Laboratories | NLM-467–1 | |
| Chemical compound, drug | D-Glucose (U-13C6, 99%; 1,2,3,4,5,6,6-D7, 97–98%) | Cambridge Isotope Laboratories | CDLM-3813–2 | |
| Chemical compound, drug | Celtone base powder (13C, 98%+; D, 97%+; 15N, 98%+) | Cambridge Isotope Laboratories | CGM-1030P-CDN-1 | |
| Chemical compound, drug | D-Glucose (U-13C6, 99%) | Cambridge Isotope Laboratories | CLM-1396–2 | |
| Chemical compound, drug | 20 Fmoc-amino acids, Fmoc-Ser(HPO3Bzl)-OH, and Fmoc-Thr(HPO3Bzl)-OH | P3Bio systems | ||
| Chemical compound, drug | Sulfo-Cy5-NHS ester | Lumiprobe | Cat. No.: 43320 | |
| Chemical compound, drug | 32P-ATP | Perkin Elmer | Cat. No.: NEG002Z2-50UC | |
| Chemical compound, drug | Pierce avidin | Thermo Scientific | Cat. No.: 21128 | |
| Chemical compound, drug | Recombinant human insulin | Thermo Scientific | Cat. No.: 12585014 | |
| Chemical compound, drug | Human insulin-like growth factor 1 (hIGF-1) | Cell Signaling Technology | Cat. No.: 8917SC | |
| Software, algorithm | ImageJ2 | DOI:10.1186/s12859-017-1934-z | ||
| Software, algorithm | GraphPad Prism version 8.2.1 | GraphPad | ||
| Software, algorithm | NmrPipe | DOI:10.1007/BF00197809 | ||
| Software, algorithm | CCPNmr Analysis version 2.4 | DOI:10.1002/prot.20449 | ||
| Software, algorithm | hmsIST | DOI:10.1007/s10858-012-9611-z | ||
| Software, algorithm | Topspin version 3.6 | Bruker |