MassIVE MSV000097370

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To study the acetylation of human serum albumin (HSA) in presence and absence of Aldicarb using 2-Naphthyl Acetate (2NA) as substrate and digestion with Pepsin.

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Description

This is study done for academic purpose and funded by ICMR, New Delhi, India, sanctioned project titled:- To study the acetylation of human serum albumin (HSA) in presence and absence of Aldicarb using 2-Naphthyl Acetate (2NA) as substrate and digestion with Pepsin. Project File No. 5/4/1-25/2020-NCD-1. Sample processing protocol: For this, 1 ml of 1 mg/ml solution of fatty acid free human albumin (SIGMA) in 10 mM Tris HCl buffer, pH 7.38 was incubated with Aldicarb (in Acetonitrile)(working concentration of 5 microMolar (Organocarbamate) for 1 hour at 37 degree Celsius. Further, acetylation was done using 2NA (prepared in Acetonitrile) for 45 minutes at 37 degree Celsius. 100 microlitre of each aliquot was denatured by boiling with 10 mM dithiothreitol for 10 minutes followed by the addition of Iodoacetamide (90 mM, SIGMA) and incubation for 30 mins in the dark at 37 degree Celsius. These were then subjected to Pepsin digestion (2 microgram, Promega, MS grade, prepared in acidic water).Next, samples to be digested by pepsin were incubated for 2 hours at 37 degree Celsius. Pepsin digestion was stopped by heating samples at 95 degree Celsius. The sample clean-up with C18 resins column was performed and the aliquots were then sent for data acquisition. Data Processing protocol: The MaxQuant software (2.0.3.1) was used for the analysis of data. Under group-specific parameters acetyl K, acetyl (N-term), and (acetyl protein N-term) were selected as variable modifications whereas the Carbamidomethyl (C) was considered a fixed modification. The label-free quantification was done and the albumin sequence file (accession: P02768) was taken as a reference for understanding the analysis. For digestion, Pepsin enzyme was added to MaxQuant enzyme file. The albumin sequence file (accession: P02768) was taken as a reference for understanding the analysis. Description of the raw files is as follow. HSA_Aldicarb is Human serum albumin treated with Aldicarb and HSA_Aldicarb_2NA is Human serum Albumin treated with Aldicarb and then substrate (2NA) was added. [doi:10.25345/C5MP4W09M] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: pesticide ; organocarbamate ; Pseudoesterase ; Albumin ; Poisoning ; DatasetType:Proteomics

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Principal Investigators:
(in alphabetical order) 		Dibyajyoti Banerjee, PGIMER, Chandigarh, India
Submitting User: SumanKaur
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