Tbc1d4

Manufactured by Cell Signaling Technology

Sourced in United States

TBC1D4 is a protein that acts as a GTPase-activating protein (GAP) for the Rab small GTPase proteins. It plays a role in the regulation of intracellular vesicle trafficking and membrane dynamics.

Automatically generated - may contain errors

Lab products found in correlation

Anti β actin monoclonal antibody, by Merck Group (1 mentions) Piperine, by Merck Group (1 mentions) Hybond electrochemiluminescence ecl nitrocellulose membrane, by GE Healthcare (1 mentions) Sb203580, by Abcam (1 mentions) Metformin, by Merck Group (1 mentions) Sodium l lactate, by Merck Group (1 mentions) Compound c, by Merck Group (1 mentions) Anti flag beads, by Merck Group (1 mentions) D 3 3h glucose, by PerkinElmer (1 mentions) Phospho akt p akt, by Cell Signaling Technology (1 mentions) Humulin r insulin, by Eli Lilly (1 mentions) High performance liquid chromatography (hplc), by Shimadzu (1 mentions) Enhanced chemi luminescence (ecl), by Thermo Fisher Scientific (1 mentions) Image lab 5, by Bio-Rad (1 mentions) Chemidoc mp, by Bio-Rad (1 mentions)

5 protocols using tbc1d4

Molecular Mechanisms of Metabolic Regulation

Check if the same lab product or an alternative is used in the 5 most similar protocols

Piperine, metformin, Sodium L-lactate, Guanosine 5′-diphosphate sodium salt (GDP, UCP1 inhibitor) and monoclonal anti-β-actin antibody were purchased from Sigma Chemical Company (St. Louis, MO, USA). Compound C (AMPK inhibitor) was provided by Merck (Rahway, NJ, USA). SB203580 (p38 MAPK inhibitor) and monoclonal antibody against UCP1 were purchased from Abcam (Cambridge, MA, USA). Monoclonal antibodies against phosphorylated AMPKα, phosphorylated p38 MAPK, phosphorylated TBC1D4, ACC, p38 MAPK, TBC1D4 and Glut4 and polyclonal antibodies against AMPKα and phosphorylated ACC were purchased from Cell Signaling Technology (Danvers, MA, USA). Monoclonal anti-c-Myc antibody was acquired from Santa Cruz Biotechnology (Dallas, TX, USA). Hybond electrochemiluminescence (ECL) nitrocellulose membranes were obtained from GE Healthcare (Little Chalfont, Buckinghamshire, UK).

Kim N., Nam M., Kang M.S., Lee J.O., Lee Y.W., Hwang G.S, & Kim H.S. (2017). Piperine regulates UCP1 through the AMPK pathway by generating intracellular lactate production in muscle cells. Scientific Reports, 7, 41066.

+ Open protocol

+ Expand

Plasmid Constructs and Antibodies

Check if the same lab product or an alternative is used in the 5 most similar protocols

The plasmid for mouse Rab10 was a generous gift from Dr. Mitsunori Fukuda. Plasmids for mouse Exoc6 and mouse Exoc6b were purchased from DNAForm and Open Biosystems, respectively. Plasmids in desired vectors were generated from these by PCR of the cDNA and ligation into appropriate restriction sites. Mutations were made by use of Agilent Technologies mutagenesis kits. All constructs were verified by DNA sequencing. An antiserum raised against a GST-fusion protein with amino acids 1–110 of mouse Exoc6 was a generous gift from Dr. Nancy C. Andrews [17 (link)]. Antibodies against phosphoserine 473 of Akt and phosphothreonine 642 of TBC1D4 were from Cell Signaling Technology, catalog numbers 9271 and 4288, respectively. Recombinant 3×Flag-tagged mouse Exoc6 and 6b were purified on anti-Flag beads (Sigma) from HEK293 cells transfected with the cDNAs for these proteins in the p3×FLAG-CMV-7.1 vector, as described in [18 (link)]. The control siRNA (catalog number 4390844) and the siRNAs for Exoc6 (catalog number 4390771, s98838) and Exoc6b were Silencer Select ones from Ambion/Life Technologies. The siRNA for Exoc6b was a custom one. Its sequence was: CCCGUUUCAAGAUAUAGAAtt.

Sano H., Peck G.R., Blachon S, & Lienhard G.E. (2015). A potential link between insulin signaling and GLUT4 translocation: association of Rab10-GTP with the exocyst subunit Exoc6/6b. Biochemical and biophysical research communications, 465(3), 601-605.

+ Open protocol

+ Expand

Rb1 Purification and Glucose Metabolism Analysis

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Rb1 purified from ginseng roots by high-performance liquid chromatography (HPLC) was purchased from Jilin University in China. HPLC (Shimadzu, Kyoto, Japan) analysis performed in our laboratory confirmed that the Rb1 had a purity of ≥98% using an Rb1 standard obtained from LKT laboratories (St. Paul, MN) (Xiong et al. 2010 (link)). d-[3-³H]glucose and 2-[¹⁴C]DG (2-deoxy-d-glucose) were purchased from PerkinElmer, Inc. (Waltham, MA). Catheters (Micro-Renathane MRE-033) were purchased from Braintree Scientific. Insulin (Humulin R) was from Eli Lilly (Indianapolis, IN), and 50% dextrose solution was from Henry Schein (Melville, NY). Antibodies for phosphorylated AMP-activated protein kinase α (pAMPKα) (Thr¹⁷²) and AMPKα, phospho-acetyl-CoA carboxylase (pACC) and ACC, GLUT4, IRS-1, phospho-Akt (pAkt), and Akt, phospho-TBC1D4 (pTBC1D4) and TBC1D4, were purchased from Cell Signaling Technology (Danvers, MA). Antibody for phospho-insulin receptor substrate 1 (pIRS-1) was purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX), and antibody for actin was from EMD Millipore (Billerica, MA).

Shen L., Haas M., Wang D.Q., May A., Lo C.C., Obici S., Tso P., Woods S.C, & Liu M. (2015). Ginsenoside Rb1 increases insulin sensitivity by activating AMP-activated protein kinase in male rats. Physiological Reports, 3(9), e12543.

+ Open protocol

+ Expand

Western Blot Analysis of Insulin Signaling

Check if the same lab product or an alternative is used in the 5 most similar protocols

10 μg of protein were resolved by SDS-PAGE and transferred to PVDF membranes. Membranes were blocked in 5% BSA or skim milk powder in TBS-T (0.1% v/v Tween 20 in Tris-buffered saline) for 1 h followed by an overnight incubation at 4 °C with specific primary antibody solutions. Membranes were incubated with an appropriate secondary antibody for 1 h before signals were detected using ECL (Thermo Scientific or Millipore) on the Chemidoc MP (Bio-Rad). Data analysis was performed using ImageJ or Image Lab version 5.2.1 (Bio-Rab). Antibodies detecting AKT, Ser(P)-473 AKT, Thr(P)-308 AKT, TBC1D4, Thr(P)-642 TBC1D4, and Tyr(P)-100 were obtained from Cell Signaling Technology. IRS-1, IRβ, TUSC5, and 14-3-3 were detected using antibodies obtained from Santa Cruz Biotechnology. Antibodies against IRAP and GLUT4 were generated in-house. Densitometry analysis was performed using Image Lab 5.2.1 (Bio-Rad). For determination of TUSC5 level, all molecular weight bands present were included for quantification.

Fazakerley D.J., Naghiloo S., Chaudhuri R., Koumanov F., Burchfield J.G., Thomas K.C., Krycer J.R., Prior M.J., Parker B.L., Murrow B.A., Stöckli J., Meoli C.C., Holman G.D, & James D.E. (2015). Proteomic Analysis of GLUT4 Storage Vesicles Reveals Tumor Suppressor Candidate 5 (TUSC5) as a Novel Regulator of Insulin Action in Adipocytes. The Journal of Biological Chemistry, 290(39), 23528-23542.

+ Open protocol

+ Expand

TBC1D4 Phosphorylation Detection

Check if the same lab product or an alternative is used in the 5 most similar protocols

TBC1D4 and phospho-TBC1D4 Ser³¹⁸, Ser⁵⁸⁸, and Thr⁶⁴² (equivalent to mouse Ser³²⁴, Ser⁵⁹⁵, Thr⁶⁴⁹) antibodies were obtained from Cell Signaling Technology. Antibodies against FLAG and HA were from Sigma-Aldrich.

Eickelschulte S., Hartwig S., Leiser B., Lehr S., Joschko V., Chokkalingam M., Chadt A, & Al-Hasani H. (2021). AKT/AMPK-mediated phosphorylation of TBC1D4 disrupts the interaction with insulin-regulated aminopeptidase. The Journal of Biological Chemistry, 296, 100637.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!