Minch, Sherrill Lynn (1996) Engineering protein glycosylation in Chinese hamster ovary cells : genetic manipulations, global glycoprotein analysis, and studies of environmental influences. Dissertation (Ph.D.), California Institute of Technology. http://resolver.caltech.edu/CaltechETD:etd-12182007-113918
The ability to engineer the glycosylation of proteins, and particularly recombinant proteins, would be of great benefit in the study and production of glycoproteins. One way to alter glycosylation is to genetically manipulate glycosyltransferase expression in a host cell. Two approaches of genetic manipulation of glycosyltransferase expression were explored: the use of a cell line with a defined alteration in the glycosylation pathway (a glycosylation "mutant") and the introduction by transfection of a new glycosyltransferase activity under the control of an inducible promoter. Additionally, the extent to which a particular genetic manipulation in the glycosylation pathway could be influenced or limited by either protein-specific effects or environmental conditions was evaluated. Optimized methods for globally surveying the response of individual glycoproteins to genetic alterations in glycosyltransferase expression were developed to aid in these evaluations. Among other results, it was demonstrated that by transfecting into a host cell the cloned glycosyltransferase [beta]-galactoside [alpha]2,6-sialyltransferase under the control of the inducible MMTV promoter, the oligosaccharides of tissue-type plasminogen activator (tPA) could be altered, under induction, to possess [alpha]2,6-linked sialic acid, a modification not detected on tPA of untransfected or uninduced cells. The methods of genetically manipulating glycosylation and evaluating the outcome of such manipulations which were explored in this report should be widely useful in efforts to tailor glycoprotein oligosaccharide structures for specific applications and to control the glycosylation of glycoproteins made in large-scale culture.
|Item Type:||Thesis (Dissertation (Ph.D.))|
|Degree Grantor:||California Institute of Technology|
|Major Option:||Chemical Engineering|
|Thesis Availability:||Restricted to Caltech community only|
|Defense Date:||28 August 1995|
|Default Usage Policy:||No commercial reproduction, distribution, display or performance rights in this work are provided.|
|Deposited By:||Imported from ETD-db|
|Deposited On:||08 Jan 2008|
|Last Modified:||26 Dec 2012 03:14|
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