Biotin-HPDP top 10

Biotin-HPDP is a sulfhydryl-reactive biotinylation reagent that forms a reversible disulfide linkage.

Biotin-HPDP is a sulfhydryl-reactive biotinylation reagent that forms a reversible disulfide linkage.Biotin-HPDP consists of a bicyclic Biotin ring structure, a 1, 6-diaminohexane attached to the side chain of biotin valonate, and a sulfhydryl reactive group at the end of the side chain[1]. It is used to label protein cysteines and other substrates that contain sulfhydryl groups.

After NEM alkylation, certain (but not all) residual free cysteine residues may react with biotin-HPDP, resulting in the coisolation of proteins containing such cysteine residues[2].

A kinetic study of the modification reactions that generate monothiophosphate disulfide linkages with either 5'-GMPS alone or 5'-GMPS-primed RNA as the substrate revealed that the second-order rate constants increased as the pH was decreased. For example, when the reaction pH was lowered from 8 to 4, the k2 value for the coupling reaction between N-(6-[biotinamido]hexyl)-3'-(2'-pyridyldithio)propionamide (biotin-HPDP) and GMPS increased 67-fold from 1.84 to 123 M(-1) x s(-1) [3].

References:

[1]: Bioconjugate Techniques , 2nd ed. By Greg T.Hermanson ?(Pierce Biotechnology, Thermo Fisher Scientific, Rockford, IL). ?Academic Press ?(an imprint of Elsevier): ?London, Amsterdam, Burlington, San Diego . 2008. ISBN 978-0-12-370501-3.

[2]: Zhou B, Wang Y, Yan Y, Mariscal J, Di Vizio D, Freeman MR, Yang W. Low-Background Acyl-Biotinyl Exchange Largely Eliminates the Coisolation of Non-S-Acylated Proteins and Enables Deep S-Acylproteomic Analysis. Anal Chem. 2019 Aug 6;91(15):9858-9866. doi: 10.1021/acs.analchem.9b01520. Epub 2019 Jul 11. PMID: 31251020; PMCID: PMC7451198.

[3]: Wu CW, Eder PS, Gopalan V, Behrman EJ. Kinetics of coupling reactions that generate monothiophosphate disulfides: implications for modification of RNAs. Bioconjug Chem. 2001 Nov-Dec;12(6):842-4. doi: 10.1021/bc0100612. PMID: 11716671.

Biotinylation method [1]:
Sample	S-nitrosylated proteins
Preparation method	Soluble in DMSO or DMF.
Reaction Conditions	50mM, 25 ℃ for 1 h
Applications	Prepare biotin-HPDP as a 50mM suspension in DMSO, Dilute with DMF to a final concentration of 4 mM. Add 1:3 volume of Labeling Solution and 1:50 volume of Ascorbate Solution（50mM） to the blocked protein samples, incubate for 1 hour at 25°C. After that, add two volumes of −20°C acetone and incubate for 20 min at −20°C to remove the biotin-HPDP. At last, Add 15 µl of packed streptavidin-agarose per mg of protein used in the initial protein sample, to purify biotinylated proteins. Incubate the biotinylated proteins with the resin for 1 hour at room temperature. Wash the beads five times with 10 volumes of Neutralization Buffer + NaCl. Centrifuge at 200g for 5 s at room temperature between each wash. Incubate the beads with Elution Buffer to recover the bound proteins. To test for the protein of interest with specific antibodies in SDS-PAGE.
References: [1].Samie R. Jaffrey and Solomon H. Snyder. The Biotin Switch Method for the Detection of S-Nitrosylated Proteins. Science’s stke.2015.

Cas No.	129179-83-5	SDF
Chemical Name	5-[(3aS,4S,6aR)-2-oxo-1,3,3a,4,6,6a-hexahydrothieno[3,4-d]imidazol-4-yl]-N-[6-[3-(pyridin-2-yldisulfanyl)propanoylamino]hexyl]pentanamide
Canonical SMILES	C1C2C(C(S1)CCCCC(=O)NCCCCCCNC(=O)CCSSC3=CC=CC=N3)NC(=O)N2
Formula	C24H37N5O3S3	M.Wt	539.78
Solubility	5.4mg/mL in DMSO (ultrasonic and warming and heat)	Storage	Store at -20°C
General tips	Please select the appropriate solvent to prepare the stock solution according to the solubility of the product in different solvents; once the solution is prepared, please store it in separate packages to avoid product failure caused by repeated freezing and thawing.Storage method and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored at -20°C, please use it within 1 month. To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time.
Shipping Condition	Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request.