Find results with:

Add another field

Search by date

to

Searching collections:

Dartmouth Dissertations

Add or remove collections

Home Dartmouth Dissertations Page 1

Reference URL

To link to the entire object, paste this link in email, IM or document

To embed the entire object, paste this HTML in website

To link to this page, paste this link in email, IM or document

To embed this page, paste this HTML in website

A novel high-cell-density recombinant protein production system based on Ralstonia eutropha.

you wish to report:

Your comment:

Your Name:

Submit

Cancel

...

Back to top

Powered by CONTENTdm®

Call No.	Ph.D. 2003
Author	Srinivasan, Sriram, 1976-
Title	A novel high-cell-density recombinant protein production system based on Ralstonia eutropha.
Date	2003
Abstract	Recombinant protein production has traditionally relied on organisms with well developed molecular biological tools, such as E. coli. However, E. coli is not suited for large industrial scale production due to its propensity to produce organic acids under oxygen-limiting conditions, inclusion body formation and proteolysis of the recombinant proteins. In this study, we describe the development of a novel protein expression system based on the industrial fermentation organism Ralstonia eutropha (formerly known as Alcaligenes eutrophus) NCIMB 40124. This new system overcomes some of the shortcomings of traditional E. coli based protein expression, in particular the propensity of inclusion body formation during high level protein expression. Using a proteomics approach we identified promoters that can be induced by manipulation of simple process parameters or media composition in high density cell culture or shake flasks respectively. By combining newly developed molecular biological tools with a high cell density fermentation process, we were able to produce high levels (1 g/liter) of soluble, active organophospho hydrolase (OPH), a model enzyme prone to inclusion body formation in E. coli. In addition, we have developed strategies based on multiple copy integration and T7 RNA polymerase-based expression to further enhance the recombinant protein titers. Using the T7 RNA polymerase approach, we have been able to produce approximately 9.5 g/liter of soluble active OPH. This represents the highest titer reported to date for this enzyme and is approximately 90 times greater than expression levels reported in E. coli.
Description	ix, 116 leaves : ill. 28 cm.
Format	application/pdf
Advisor	Tillman U. Gerngross Lee R. Lynd Stefan Wildt David W. Wood.
Subject	Recombinant proteins -- Synthesis. Ralstonia. Alcaligenes eutrophus. Organophosphorus compounds -- Synthesis. Proteomics. Microbial biotechnology. Industrial microbiology. Recombinant Proteins -- Biosynthesis.
College	Thayer School of Engineering.
Issuing Body	Dartmouth Dissertations are made available through the Dartmouth Digital Library.
Degree	Thesis (Ph.D.)--Dartmouth College.
Copyright	All rights reserved. No part of this book may be reproduced in any form by any electronic or mechanical means without permission in writing from the author.
DOI	http://dx.doi.org/10.1349/ddlp.31

Title	Page 1
Format	application/pdf
Issuing Body	Dartmouth Dissertations are made available through the Dartmouth Digital Library.
Degree	Thesis (Ph.D.)--Dartmouth College.
Full Text	A Novel High-cell-density Recombinant Protein Production System Based on Ralstonia eutropha A Thesis Submitted to the Faculty In partial fulfillment of the requirements for the degree of Doctor of Philosophy By Sriram Srinivasan Thayer School of Engineering Dartmouth College Hanover, New Hampshire July 2003

Find results with:

Searching collections:

Home Dartmouth Dissertations Page 1

A novel high-cell-density recombinant protein production system based on Ralstonia eutropha.

Object Description

Description

A novel high-cell-density recombinant protein production...

A novel high-cell-density recombinant protein production...

	All
	Pages Enter page numbers or page ranges separated by commas. For example: 1,3,5-12