Chemical and physical properties of bovine alpha-lactalbumin, beta-casein and genetically altered bovine beta-casein produced in the milk of transgenic mice

The genomic bovine α-lactalbumin gene (1.7 kb) and the bovine β-casein gene (8.5 kb) linked to bovine α-lactalbumin 5$\prime$ flanking region (2.0 kb) were expressed in two lines of transgenic mice, α-114 and αβ 34, respectively. Whey proteins, caseins, or skim milk proteins of the transgenic mouse milk containing bovine α-lactalbumin or bovine β-casein were analyzed and separated by polyacrylamide gel electrophoresis (PAGE), using SDS, urea, or native (non-denaturing) conditions. Using PAGE followed by immunoblotting, both heterologous proteins were proved as identical as the native proteins in the molecular weight and the charge. The estimated expression level of bovine α-lactalbumin and β-casein was about 1.0 mg/ml and 3.0 mg/ml, respectively. The expression level of bovine α-lactalbumin was lactation-dependent, which was higher in early lactation than that in late lactation stage. Calcium binding of the heterologous bovine α-lactalbumin was also the same as native bovine α-lactalbumin. The phosphorylation level of the bovine β-casein expressed in the transgenic mouse milk was identical to the native bovine β-casein. The glycosylated bovine α-lactalbumin and the N-terminal amino acid sequence of purified bovine α-lactalbumin were characterized and determined. The physical properties of transgenic mouse milks were changed due to the heterologous expression of the bovine β-casein or α-lactalbumin in the mouse milks. An increased moisture of transgenic mouse milk containing bovine α-lactalbumin was observed. However, the milk composition of transgenic mouse milk containing bovine β-casein did not change significantly. In addition, two chymosin-resistant and one plasmin-resistant bovine β-casein genomic constructs were generated and microinjected to generate transgenic mice. Mouse milk containing each of the three mutant β-caseins was produced and analyzed by immunoblotting with a bovine β-casein specific antibody. The mice carrying one of the chymosin-resistant gene constructs produced the milk containing mutant bovine β-casein with the expected molecular weight. The result of rennin digestion demonstrated that the peptide bond of 192-193 in the mutant protein was chymosin-resistant. However, the mice carrying the plasmin-resistant β-casein produced two proteins in their milk which reacted with bovine β-casein antibody and had smaller molecular weights than that of the expected mutant β-casein.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Food Science and Human Nutrition

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2011

Author dc:creator

• Jeng, Shin-Yi

Contributors dc:contributor

• Jimenez-Flores, Rafael

• Agriculture, Food Science and Technology
• Biology, Molecular

Statement dc:rights

• Copyright 1996 Jeng, Shin-Yi

Language dc:language

eng

Identifier

9780591087307
AAI9702549
(UMI)AAI9702549

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/19207