Some interesting facts 
- A tetramer of 4 identical subunits
- Each subunit is 120kD.
- Active only as a tetramer.
- Mutations in some of the codons of the N-terminal 60aa or C-terminal 100aa results in an inactive, dimeric β-galactosidase. 
- If the above sequences are deleted, the missing protein fragment can be replaced by the corresponding peptide. This is called intracistronic alpha or omega complementation respectively.
- Its N-terminal 23 residues can be replaced by any amino acid residues without affecting the enzymatic activity.
- A mutant with an internal deletion of codons 21-41 of the lacZ gene does not produce any active β-galactosidase.
- A mutant with a deletion of everything past residue 60 (i.e. it expresses only the first 60 N-terminal amino acids) does not produce any active β-galactosidase.
- You can measure lacZ activity using flow cytometry. See A flow cytometric study of stationary phase gene expression in E.coli using lacZ reporter gene fusion
- Alpha-complementation of β-galactosidase does not seem to yield activities equal to wildtype β-galactosidase. Depending on the fragment, the activity can be up to 24% of wildtype . (If anyone has a better reference comparing results from a Miller assay of alpha-complementated β-galactosidase with wildtype, please include it here.)
- Beta-Galactosidase Assay (A better Miller)
- BE.109:Protein engineering/Assessing beta-galactosidase
- See also Beta-galactosidase assays in yeast
- Plovins A, Alvarez AM, Ibañez M, Molina M, and Nombela C. Use of fluorescein-di-beta-D-galactopyranoside (FDG) and C12-FDG as substrates for beta-galactosidase detection by flow cytometry in animal, bacterial, and yeast cells. Appl Environ Microbiol. 1994 Dec;60(12):4638-41.
- Zamenhof PJ and Villarejo M. Construction and properties of Escherichia coli strains exhibiting -complementation of -galactosidase fragments in vivo. J Bacteriol. 1972 Apr;110(1):171-8.
- Ullmann A. Complementation in beta-galactosidase: from protein structure to genetic engineering. Bioessays. 1992 Mar;14(3):201-5. DOI:10.1002/bies.950140311 |
- [by] Jeffrey H. Miller. Experiments in molecular genetics. [Cold Spring Harbor, N.Y.] Cold Spring Harbor Laboratory, 1972. ISBN:0879691069
original β-galactosidase assay by Miller
- Thibodeau SA, Fang R, and Joung JK. High-throughput beta-galactosidase assay for bacterial cell-based reporter systems. Biotechniques. 2004 Mar;36(3):410-5.
- Serebriiskii IG and Golemis EA. Uses of lacZ to study gene function: evaluation of beta-galactosidase assays employed in the yeast two-hybrid system. Anal Biochem. 2000 Oct 1;285(1):1-15. DOI:10.1006/abio.2000.4672 |