Understanding the biochemical properties and physiological function of the protein syncollin
type: CreativeWork
description: Syncollin is a 16-kDa protein that was originally isolated from the pancreatic zymogen granule. It is now known also to be expressed in the gut, the spleen and in neutrophils. The protein contains intramolecular disulphide bonds and is present both free within the ZG lumen and tightly associated with the luminal leaflet of the ZG membrane; it is also secreted into the pancreatic juice. Syncollin is able to oligomerize, and assemble into doughnut-shaped structures, which might explain its known pore-forming activity. Syncollin appears to be involved in a number of gut-based disease states. For instance, syncollin expression was found to be down-regulated in the colon when a bacterial suspension was administered to germ-free mice, and in mice with chemically-induced colitis-associated cancer. The available evidence suggests that syncollin plays an important role in the gut, and possibly elsewhere. This dissertation describes my attempts to understand this role and its structural basis. I first assessed various methods for purification of recombinant syncollin. Syncollin was expressed with various epitope tags (including His, GST and Strep) in bacteria, insect cells and mammalian cells. The best results were obtained by expressing syncollin bearing a double-Strep tag at its C terminus (syncollin-Strep) in mammalian (tsA-201) cells and purifying the protein from the cell supernatant using the Strep-Tactin XTTM system. Syncollin-Strep purified in this way contained intra-molecular disulphide bonds and recapitulated the ability of the native protein to bind to syntaxin 2 and permeabilize membranes. In the pancreatic juice, syncollin will encounter an environment rich in proteolytic activity. One might expect, therefore, that its structure would be highly stable. To test this hypothesis, I assessed the thermal stability of the protein using circular dichroism (CD) spectroscopy. The CD spectrum of syncollin-Strep indicated a predominantly beta-sheet structure. When the prot…
abstract: Syncollin is a 16-kDa protein that was originally isolated from the pancreatic zymogen granule. It is now known also to be expressed in the gut, the spleen and in neutrophils. The protein contains intramolecular disulphide bonds and is present both free within the ZG lumen and tightly associated with the luminal leaflet of the ZG membrane; it is also secreted into the pancreatic juice. Syncollin is able to oligomerize, and assemble into doughnut-shaped structures, which might explain its known pore-forming activity. Syncollin appears to be involved in a number of gut-based disease states. For instance, syncollin expression was found to be down-regulated in the colon when a bacterial suspension was administered to germ-free mice, and in mice with chemically-induced colitis-associated cancer. The available evidence suggests that syncollin plays an important role in the gut, and possibly elsewhere. This dissertation describes my attempts to understand this role and its structural basis. I first assessed various methods for purification of recombinant syncollin. Syncollin was expressed with various epitope tags (including His, GST and Strep) in bacteria, insect cells and mammalian cells. The best results were obtained by expressing syncollin bearing a double-Strep tag at its C terminus (syncollin-Strep) in mammalian (tsA-201) cells and purifying the protein from the cell supernatant using the Strep-Tactin XTTM system. Syncollin-Strep purified in this way contained intra-molecular disulphide bonds and recapitulated the ability of the native protein to bind to syntaxin 2 and permeabilize membranes. In the pancreatic juice, syncollin will encounter an environment rich in proteolytic activity. One might expect, therefore, that its structure would be highly stable. To test this hypothesis, I assessed the thermal stability of the protein using circular dichroism (CD) spectroscopy. The CD spectrum of syncollin-Strep indicated a predominantly beta-sheet structure. When the prot…
author: Waters, Rosie
language: eng
keywords: Antimicrobial peptide, Host defence, Pancreatic zymogen granule, Membrane permeabilisation, Recombinant protein purification
license: https://www.rioxx.net/licenses/all-rights-reserved/
date published: 2021-07-30
date modified: 2026-07-22T22:24:30Z
provider: Cambridge University
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Cambridge

PropertyValue
property ID: DOI
value: 10.17863/CAM.79580