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Activity of Recombinant Cysteine-Rich Domain Proteins Derived From the Membrane-Bound MUC17/Muc3 Family Mucins

Activity of Recombinant Cysteine-Rich Domain Proteins Derived From the Membrane-Bound MUC17/Muc3 Family Mucins

Samuel B Ho, Ying Luu, Laurie L Shekels, Surinder K Batra, Brandon Kandarian, David B Evans, Phillip G Zaworski, Cindy L Wolfe, Robert L Heinrikson

Biochim Biophys Acta. 2010 Jul;1800(7):629-38.

PMID: 20332014

Abstract:

Background:
The membrane-bound mucins, MUC17 (human) and Muc3 (mouse), are highly expressed on the apical surface of intestinal epithelia and have cytoprotective properties. Their extracellular regions contain two EGF-like Cys-rich domains (CRD1 and CRD2) connected by an intervening linker segment with SEA module (L), and may function to stimulate intestinal cell restitution. The purpose of this study was to determine the effect of size, recombinant host source, and external tags on mucin CRD1-L-CRD2 protein activity.
Methods:
Four recombinant Muc3-CRD proteins and three MUC17-CRD proteins were generated using Escherichiacoli or baculovirus-insect cell systems and tested in colonic cell cultures for activity related to cell migration and apoptosis.
Results:
N-terminal glutathione-S-transferase (GST) or C-terminal His(8) tags had no effect on either the cell migration or anti-apoptosis activity of Muc3-CRD1-L-CRD2. His-tagged Muc3-CRD1-L-CRD2 proteins with truncated linker regions, or the linker region alone, did not demonstrate biologic activity. The human recombinant MUC17-CRD1-L-CRD2-His(8) was shown to have anti-apoptotic and pro-migratory activity, but did not stimulate cell proliferation. This protein showed similar in vitro biologic activity, whether produced in E. coli or a baculovirus-insect cell system.
Conclusions:
Recombinant mucin proteins containing a bivalent display of Cys-rich domains accelerate colon cell migration and inhibit apoptosis, require a full-length intervening Linker-SEA segment for optimal biologic activity, and are functional when synthesized in either E. coli and insect cell systems.
General significance:
These results indicate that an Escherichiacoli-derived full-length His(8)-tagged human MUC17 CRD1-L-CRD2 recombinant protein is a biologically active candidate for further development as a therapeutic agent.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
IAR42411820	CAMK1δ, active, GST tagged human			Price
IAR42416204	p70S6Kb, active, GST tagged human			Price

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