Expression and bioactivity of recombinant human lysozyme in the milk of transgenic mice.
Human milk lysozyme is an important protein for innate immunity, but human breast milk is a fairly poor source for commercial production of this enzyme. Research on the expression of recombinant human lysozyme (rHlys) is therefore potentially valuable to the dairy industry. In this study, 2 different kinds of transgenic mice, PBC-hLY and PBC-sighLY, were generated and used as system models to express rHlys. Six lines of PBC-hLY transgenic mice with human lysozyme genomic DNA-based constructs were generated, and a maximum expression level of rHlys approaching 0.154 mg/mL was achieved. Antibacterial activity of the whey from PBC-hLY female transgenic mice was determined by a turbidimetric assay. Results showed that antibacterial activity of the whey was strongly enhanced, and confirmed that rHlys retained full activity. For rHlys to be secreted efficiently into the milk of transgenic mice, 5 lines of mice were also generated, in which the signal peptide DNA of bovine beta-casein was substituted for that of lysozyme in PBC-hLY transgenic mice. Compared with PBC-hLY transgenic mice, both the expression levels of rHlys and the antibacterial activity of the whey were much higher in the PBC-sighLY transgenic mice. The concentration of rHlys in one of these mice amounted to 1.405 mg/mL-3 times higher than the level in human whey. The antibacterial activity of the whey was also 3 times higher than that of human whey. The rHlys from both PBC-hLY and PBC-sighLY transgenic mice had the same antibacterial activity as human milk lysozyme. The effect of the signal peptide and copy numbers of the transgene on expression of rHlys was also evaluated. This work will certainly permit a better understanding of how mammary gland bioreactor systems can be applied to produce rHlys in other mammals, such as cattle.
Yu
Z
Z
State Key Laboratory for Agrobiotechnology, China Agricultural University, Beijing 100094, People's Republic of China.
Meng
Q
Q
Yu
H
H
Fan
B
B
Yu
S
S
Fei
J
J
Wang
L
L
Dai
Y
Y
Li
N
N
D000818:Animals; D015139:Blotting, Southern; D015153:Blotting, Western; D002473:Cell Wall / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D008837:Micrococcus / Q000648:ultrastructure; D008895:Milk, Human / Q000201:enzymology; D009113:Muramidase / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins; D014162:Transfection
D011994:Recombinant Proteins; D009113:Muramidase