Identification of sites of N-glycosylation of human and mouse Tamm-Horsfall glycoprotein with the aid of database searching

Koe, Chun Iq (2006) Identification of sites of N-glycosylation of human and mouse Tamm-Horsfall glycoprotein with the aid of database searching. Universiti Malaysia Sabah. (Unpublished)

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Abstract

A potential N-glycosylation site can be either glycosylated or unglycosylated or partially glycosylated. Therefore, it is important to identify the N-glycosylation sites of a glycoprotein in order to understand its function and character. Tamm-Horsfall glycoprotein is the most abundant protein in mammalian urine. The human and mouse Tamm-Horsfall glycoproteins reasonably pure were isolated using Tamm and Horsfall method. The isolation method can isolate Tamm-Horsfall glycoprotein with single band for each sample (HMl, HFl, HF2, and MFl) on SDS-PAGE. All human samples (HMl, HFI and HF2), have identical size around 100-120 kDa in reduced form. The mouse sample (MFl) has heavier size (around 115-120 kDa) than the human sample. Bradford protein assay was done on a human sample (HMl) and showed that there is 0.0272 milligrams of human Tamm-Horsfall glycoproteins in every millilitre of 0.4 mg/mL of protein solution. The database searching is done to identify the site of N-glycosylation. The data of human and mouse Tamm-Horsfall glycoproteins (including precursor) were retrieved from Swiss-Prot database. From the Swiss-Prot database, the expected region of human Tamm-Horsfall glycoprotein (including precursor) to be seen on a human kidney 2D-PAGE was predicted using SWISS-2DPAGE based on the pI and molecular weight computed by the database. The mass of peptide fragments of human and mouse Tamm-Horsfall glycoproteins generated by trypsin with carboxymethylation were computed using PeptideMass. Both human and mouse Tamm-Horsfall glycoproteins N-glycosylation sites were identified using NetNGlyc. Human Tamm-Horsfall glycoprotein shows total eight N-glycosylation sites (Asn³�, Asn��, Asn�º, Asn²³², Asn²��, Asn³²², Asn3��, and Asn�¹�) with all sites predicted to be glycosylated except Asn³��. Mouse Tamm-Horsfall glycoprotein shows total ten N-glycosylation sites (Asn²�, Asn³�, Asn��, Asn��, Asn²³³, Asn���, Asn³²³, Asn³��, Asn���, and Asn�¹�) with all ten sites predicted to be glycosylated.

Item Type: Academic Exercise
Uncontrolled Keywords: N-glycosylation, isolation method, Tamm-Horsfall, Swiss-Prot database, mammalian urine, peptide fragment, trypsin, carboxymethylation, molecular weight
Subjects: Q Science > QH Natural history > QH301 Biology
Divisions: SCHOOL > School of Science and Technology
Depositing User: MDM SITI AZIZAH IDRIS
Date Deposited: 08 Apr 2014 03:44
Last Modified: 13 Oct 2017 02:24
URI: http://eprints.ums.edu.my/id/eprint/8693

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