Downstream processing of recombinant and endogenous proteins from livestock milk

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.

Name thesis:degree_name

Ph. D.

Level thesis:degree_level

doctoral

Discipline thesis:degree_discipline

Chemical Engineering

Department dc:contributor.department

Chemical Engineering

Grantor dc:publisher

Virginia Tech

Year dc:date.issued

1999

Author dc:creator

• Degener, Arthur W. Jr.

Chair dc:contributor.committeechair

• Velander, William H.

Committee members dc:contributor.committeemember

• Drohan, William N.
• Williams, Kimberly Forsten
• Conger, William L.
• Davis, Richey M.

• endogenous milk proteins
• recombinant proteins
• free flow iso-electric focusing
• Free flow electrophoresis
• Expanded bed adsorption chromatography

Statement dc:rights

• In Copyright

Licence dc:rights.uri

• http://rightsstatements.org/vocab/InC/1.0/

Dc Identifier Other

etd-042399-164339

OAI identifier oai:identifier

oai:vtechworks.lib.vt.edu:10919/27183