Title page for ETD etd-042399-164339


Type of Document Dissertation
Author Degener, Jr., Arthur W.
Author's Email Address adegener@vt.edu
URN etd-042399-164339
Title Downstream processing of recombinant and endogenous proteins from livestock milk
Degree PhD
Department Chemical Engineering
Advisory Committee
Advisor Name Title
Velander, William H. Committee Chair
Conger, William L. Committee Member
Davis, Richey M. Committee Member
Drohan, William N. Committee Member
Williams, Kimberly Forsten Committee Member
Keywords
  • endogenous milk proteins
  • recombinant proteins
  • free flow iso-electric focuing
  • Free flow electrophoresis
  • Expanded bed adsorption chromatography
Date of Defense 1999-04-08
Availability unrestricted
Abstract
With the increased demands of therapeutic proteins, there is going to be a need for new purification technologies which have high throughput, high yield and high resolution. Three purification technologies were explored as potential new technology to isolate recombinant and endogenous milk proteins: Expanded bed adsorption chromatography(EBAC) combined with hydrophobic interaction chromatography(HIC), Recycle continuous flow electrophoresis(RCFE) and Free flow isoelectric focusing(FFIEF). The first process(EBAC/HIC) used with Zn2+ as a selective precipitating agent, purified recombinant human protein C(rhPC) and IgG(contaminated with less than 1% IgA) from swine milk with high resolution and high yield while processing about 10-20 grams in a single operation. The second process(RCFE) was able to isolate the active sub-populations of rhPC from major milk contaminants( - and -pig casein) as wells as from the inactive sub-populations of rhPC. RCFE was able to process 1.5g total protein per hour on a small scale and is currently being researched to process 1kg total protein per hour. The third and final purification process(FFIEF) sub-fractionated 100mg of immuno-purified rhPC into 50 fractions. The FFIEF was able to produce a linear pH gradient over the range of 3-10 using 2% ampholytes. The fractionated rhPC showed differing degrees of activity that resulted from the -carboxylated glutamic acids and the sialic acids.

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