a handle used to describe a protein identified by an antibody to a specific acetylation site
a modification used to represent acetylation
activated; used when a protein has been shown experimentally to be capable of catalyzing a reaction
a modification used to represent activation
a handle used to describe a protein or chemical immoblized on agarose
molecules containing an amine group, a carboxylic acid group and a side chain that varies between different amino acids.
a Treatment used when mixtures of amino acids are added to cells
used when the substrate of a kinase reaction is the kinase itself
DNA in the B conformation
represented by Poly(dA-dT)-Poly(dT-dA) 17618271(M) 16785313(M)
the right-handed typical form of double helix DNA in which the chains twist up and to the right around the front of the axis of the helix and that has usually ten base pairs in each helical turn and two grooves on the external surface [Merriam-Webster Dictionary]
an ELISA performed on suspended beads and analysed by FACS
sample containing bGal is incubated with colorless substrate ONPG (o-nitrophenyl-beta-D-galactopyranoside). Active enzyme hydrolyzes subtrate to o-nitrophenyl which is yellow. The reaction is terminated with sodium carbonate and the absorbance at 420 nM is measured by spectrophotometry. [Promega catalog]
bimolecular ﬂuorescence complementation
A method of viewing the association of proteins inside living cells. The intact Green fluorescent protein (and its variants CFP, YFP, BFP, and RFP) is fluorescent. However, when the fluorescent protein is split into N and C-terminal halves, the molecule does not produce fluorescence. Fusing of each of the two non-fluorescent fragments to two putative interacting partners leads to restoration of fluorescence within a cell by reconstituting the split fluorophore. This fluorescence is detected via fluorescence microscopy, which can be recorded by a mounted camera. The advantage of the BiFC method over other methods of visualizing protein-protein interactions is that it gives an indication of interaction, as well as cellular localization of the complex. [Wiki]
Hu CD, Chinenov Y, Kerppola TK (2002) Visualization of interactions among bZIP and Rel family proteins in living cells using bimolecular ﬂuorescence complementation. Mol Cell 9: 789-798
Hu CD, Grinberg A, Kerppola T (2005) Visualization of protein interaction in living cells using bimolecular ﬂuorescence complementation (BiFC) analysis. In Current Protocol in Cell Biology, Bonifacino JS, Dasso M, Harford JB, Lippincott-Schwartz J, Yamada KM (eds), pp 21.3.1–21.3.21. Hoboken: John Wiley & Sons
Hu CD, Kerppola TK (2003) Simultaneous visualization of multiple protein interactions in living cells using multicolor ﬂuorescence complementation analysis. Nat Biotechnol 21: 539–545
a streptavidin binding protein used as a protein tag
a handle used to describe a protein or chemical prebound to biotin
a detection method used for detecting protein surface expression
DRM and non-DRM fractions were separated as described previously (Shima et al, 2003). Briefly, cells were lysed in buffer A (50 mM Tris–HCl (pH 7.4), 150 mM NaCl, 1 mM EDTA, 1 mM sodium orthovanadate, 50 mM NaF, 5 mM 2-mercaptoethanol and protease inhibitors) containing 0.25% Triton X-100 at 41oC. The lysate was separated on a discontinuous sucrose gradient (40–35–5%) by ultracentrifugation at 100000g for 16h at 4oC. The gradient was separated into 12 fractions collected from the top. The DRM proteins were solubilized in buffer A containing 2% N-octyl-b-D-glucoside (ODG) and 1% Nonidet P40 (NP40)
To analyze cellular responses to DSBs, including repair, at defined sites in human cells, we used the eukaryotic homing endonuclease I-PpoI, which has a 15 base pair recognition sequence7,8, to cleave endogenous DNA target sites in the human genome. Expression of I-PpoI in human cells results in cleavage of ~10% of the 200–300 I-PpoI genomic target sites9 to generate ~30 DSBs per cell (equivalent to that introduced by exposure to ~0.8 Gy irradiation).
The system was improved by adding a mutant oestrogen receptor hormone-binding domain to I-PpoI to create a fusion protein that could be localized to the nucleus in response to 4-hydroxytamoxifen (4-OHT). Addition of 4-OHT to MCF7 cells infected with an oestrogen receptor–I-PpoI retrovirus resulted in time-dependent cleavage of the endogenous 28S rDNA I- PpoI site and ATM activation.
double stranded DNA
double stranded RNA
transcribed in vitro from PCR products using T7 RNA polymerase 16625202(M)
a handle used to enable activated transcription factors to interact with a Gal4-reporter
a string of nucleotides that codes for a Protein
Names of Genes in this model are derived from the Protein name (eg. the name of the gene for the protein Erk1 is Erk1-gene)
used as a GTP analog by 16892055
used as a GTP analog by 9001246
CID 36735 Guanylyl Imidodiphosphate
A non-hydrolyzable analog of GTP, in which the oxygen atom bridging the beta to the gamma phosphate is replaced by a nitrogen atom. It binds tightly to G-protein in the presence of Mg2+. The nucleotide is a potent stimulator of adenylate cyclase. [PubChem]
a handle used to describe or isolate a protein fused to GST
nonhydrolyzable G-protein-activating analog of GTP
a Sort used in curation shorthand to describe the way a protein is identified in an experiment
knockdown in a cell line by homologous recombination
two promoterless targeting vectors containing either a geneticin or hygromycin resistance gene in place of genomic p53 sequences were sequentially transfected into HCT116 p53+/+ cells to disrupt both p53 alleles  18324520
. Bunz F, Dutriaux A, Lengauer C, Waldman T, Zhou S, Brown JP, Sedivy JM, Kinzler KW, Vogelstein B. Requirement for p53 and p21 to sustain G2 arrest after DNA damage. Science 1998;282:1497–1501. [PubMed: 9822382]
a prefix used to designate a protein expressed in insect cells and purified by immunoprecipitation or pulldown
Inductively coupled plasma atomic emission spectroscopy (ICP-AES), also referred to as inductively coupled plasma optical emission spectrometry (ICP-OES), is an analytical technique used for the detection of trace metals. It is a type of emission spectroscopy that uses the inductively coupled plasma to produce excited atoms and ions that emit electromagnetic radiation at wavelengths characteristic of a particular element. The intensity of this emission is indicative of the concentration of the element within the sample. [Wikipedia]
In Cell Western, a detection method
Treated cells are:
fixed by the addition of 50 μL of a 5X Mirsky’s solution (National Diagnostics, Atlanta, GA) for 60 min
washed in Tris-buffered saline, pH 7.4.
incubated in TBS containing 0.1% Triton-X100 and 0.1% bovine serum albumin (blocking buffer) for 2 h
incubated overnight at 4°C with primary antibodies diluted in blocking buffer
washed in TBS
incubated with europium-labeled secondary antibody for 90 min at room temperature in the dark
washed in TBS
incubated with Delphia Enhancement Solution for 120 min
Fluorescence is quantified an Envision 2101 multilabel reader (Perkin Elmer, Waltham, MA).
The complex isolated from a lysate using an antibody against a protein or a protein-tag, or by pull down with things other than antibodies the bind to protein or protein-tag (such as GST:GSH, m7GTP, biotin:streptavidin)
the source of an immunoprecipite used in a test-tube experiment
in response to
used when cells are treated with an external agent (addition of a reagent or application of a stress)
in vitro kinase assay
an assay used to determine the ability of the complex immunoprecipitated with a protein to phosphorylate a substrate
7-methyl-GTP Sepharose, a synthetic analog of the mRNA 7-methylguanosine cap complex
a handle used to represent capped-mRNA
Maltose binding protein
a handle used to describe or isolate a protein fused to MalBP
Mammalian Two Hybrid method
In the mammalian two-hybrid system, protein-protein interactions are detected by fusing one test protein to the Gal4 DNA binding domain, fusing a second test protein to the strong transcriptional activation domain of the VP16 protein, and coexpressing these with the GAL4-dependent reporter. An interaction between these two proteins results in activation of a Gal4-dependent luciferase reporter by the VP16 activation domain.
a handle used when a protein is identified by Mass Spectroscopy
bacteriophage MS2 coat protein
a handle used to describe a protein fused to MS2
a change in the sequence of a protein
a Sort used in curation shorthand to describe the mutation of a protein
a handle used when a protein is identified by its migration on an SDS-PAGE gel
a mutation that targets proteins to the cell membrane
note: when myr is used as a mutation it means that a myristolyation site (usually from Src) is fused at the N-terminus of an expressed protein
Native Western Blot
Proteins are separated by gel electrophoresis under non-denaturing conditions. The proteins are then transferred to a membrane (typically nitrocellulose or PVDF), where they are detected using antibodies specific to the target protein.
Lysates are run on non-denaturing PAGE gels, blotted, and stained for Subject.
Note: If the dectected Protein complex is approximately twice the molecular weight of the uncomplexed Protein, then the assay is called "dimerization". Otherwise, "oligomerization" is used.
Limitations: Does not distinquish between a complexes of Subject:Subject and Subject:OtherProtein (of a similar size).
Selective amplification via biotin- and restriction-mediated enrichment
Lavery,D.J., Lopez Molina,L., Fleury,O.F. and Schibler,U. (1997) Selective amplification via biotin- and restriction-mediated enrichment (SABRE), a novel selective amplification procedure for detection of differentially expressed mRNAs. Proc. Natl Acad. Sci. USA, 94, 6831–6836.
Denatured proteins are separated using SDS-PAGE. The proteins are then transferred to a membrane (typically nitrocellulose or PVDF), where they are detected using antibodies specific to the target protein.