methyl β cyclodextrin complexed cholesterol  (MedChemExpress)

Journal: The Journal of Biological Chemistry

Article Title: The mammalian cholesterol synthesis enzyme squalene monooxygenase is proteasomally truncated to a constitutively active form

doi: 10.1016/j.jbc.2021.100731

Figure Lengend Snippet: A truncated and cholesterol- resistant form of SM is present in a variety of cell types. A , the indicated cell lines were treated in the presence or the absence of 1 μM NB-598 for 8 h, and immunoblotting was performed for SM and truncated SM (trunSM; red ). Immunoblots are representative of n ≥ 3 (HEK293T, HeLaT, Huh7, and HepG2) or n = 1 (Be(2)-C) independent experiments. B and C , HEK293T cells were treated with 10 μg/ml cycloheximide in the presence or the absence of ( B ) 1 μM NB-598 or ( C ) 20 μg/ml cholesterol-methyl-β-cyclodextrin complexes (Chol/CD) for the indicated times. Graphs depict densitometric quantification of SM levels ( left ), trunSM levels ( center ), or total SM levels (SM + trunSM; right ) normalized to the 0 h time point, which was set to 1 ( dotted line ). Data presented as mean ± SEM from n ≥ 3 independent experiments (∗ p ≤ 0.05; two-tailed paired t test versus vehicle condition). HEK293T, human embryonic kidney 293T; SM, squalene monooxygenase.

Article Snippet: Chemicals were from the following suppliers: ALLN (A6185; Sigma–Aldrich), ammonium chloride (31; Ajax Finechem), l -azidohomoalanine (1066; Click Chemistry Tools), bafilomycin A1 (B1793; Sigma–Aldrich), β-mercaptoethanol (M3148; Sigma–Aldrich), bromophenol blue (B0126; Sigma–Aldrich), butylated hydroxytoluene (B1378; Sigma–Aldrich), [ 14 C]-acetate sodium salt (PerkinElmer; catalog no. NEC084H001MC), CB-5083 (16276; Cayman Chemical Company), cholesterol complexed with methyl-β-cyclodextrin (Chol/CD; C4951; Sigma–Aldrich), chloroform (102444; Supelco), cycloheximide (C7698; Sigma–Aldrich), l -cysteine (168149; Sigma–Aldrich), diethyl ether (code: EA036; Chem-Supply), dimethyl sulfoxide (DMSO) (2225; Ajax Finechem), ethanol (214; Ajax Finechem), glacial acetic acid (100063; Supelco), GlutaMAX (M4667; Thermo Fisher), glycerol (242; Ajax Finechem), glycine (1083; Ajax Finechem), Hepes (54457; Sigma–Aldrich), heptane (LC1078; RCI LabScan), hexane (104369; Supelco), hydrochloric acid (256; Ajax Finechem), IGEPAL CA-630 (I8896; Sigma–Aldrich), magnesium chloride (296; Ajax Finechem), methanol (318; Ajax Finechem), l -methionine (M9625; Sigma–Aldrich), methotrexate (13960; Cayman Chemical Company), mevalonolactone (mevalonate; M4667; Sigma–Aldrich), mevastatin (compactin; M2537; Sigma–Aldrich), MG132 (C2211; Sigma–Aldrich), NB-598 (CS-1274; Chemscene), PEG4 carboxamide-propargyl biotin (1266; Click Chemistry Tools), polyethyleneimine (03880; Sigma–Aldrich), Ponceau S solution (P7170; Sigma–Aldrich), potassium chloride (383; Ajax Finechem), potassium hydroxide (405; Ajax Finechem), PR-619 (16276; Cayman Chemical Company), Ro 48-8071 (10006415; Cayman Chemical Company), sodium carbonate (463; Ajax Finechem), sodium chloride (465; Ajax Finechem), sodium deoxycholate (D6750; Sigma–Aldrich), SDS (75746; Sigma–Aldrich), sodium EDTA (180; Ajax Finechem), sodium EGTA (E8145; Sigma–Aldrich), sodium hydroxide (482; Ajax FineChem), sodium orthovanadate (S6508; Sigma–Aldrich), sodium pyruvate (11360070; Thermo Fisher), Tris (2311; Ajax Finechem), Tween-20 (P9416; Sigma–Aldrich), and WP1130 (15277; Cayman Chemical Company).

Techniques: Western Blot, Two Tailed Test