This is about the enzyme beta galactosidase. You will know about the complete information of this enzyme.

1. DNA Marker/Reporter Enzyme
β-galactosidase
Presented by : Fakeh Iqbal
Roll No:424 -M.phil-(M)Z-21

2. DNA MARKER
A genetic marker/ DNA marker is a DNA sequence with a specific
location on a chromosome.
Genetic markers help in various ways as its link an inherited disease
with the responsible gene.
DNA segments which are repeated again and again in DNA.
Genetic markers are used to track the inheritance of different genes
whose approximate location is known.

4. β-Galactosidase
The enzyme beta-galactosidase was first mentioned in the literature
by Beijerinck exactly a hundred years ago.
β-galactosidase is also called lactase, beta-gal or β-gal belongs to the
family of glycoside hydrolase enzymes.
It catalyzes the hydrolysis of β galactosides into monosaccharides
through the breakage of glycosidic bond.
β-galactosides include carbohydrates containing galactose where the
glycosidic bond present above the galactose molecule.

5. Substrates of β-Galactosidase
Ganglioside GM1
Lactosylceramides
Lactose
Various glycoproteins

6. Inhibitors of β-galactosidase
Beta-gal is inhibited by L-ribose,
Non-competitive inhibitor iodine,
Competitive inhibitors
 2-phenylethyl 1-thio-beta-D-galactopyranoside (PETG),
 D-galactonolactone,
 isopropyl thio-beta-D-galactoside (IPTG),
 galactose

7. β-Galactosidase Assay
β-Gal as a reporter gene
Protein product is stable and resistant to proteolytic degradation.
Enzyme is easily assayed.
Easy and rapid method to assay β-gal activity in transfected cells.
No expensive equipment.

9. Families of β-galactosidase
Most of the work done on β-galactosidase is derived from E. coli. However
β-gal can be found in many plants especially in fruits, mammals, yeast,
bacteria, and fungi.
β-galactosidase genes may be differ in the length of their coding sequence
and the length of proteins formed by amino acids.
According to the sequence the β-galactosidase is divided into four families:
GHF-1 (Glycoside hydrolase)
GHF-2
GHF-35
GHF- 42

10. Conti ….
E. Coli belongs to GHF-2, all plants belong to GHF-35,
and Thermus thermophilus (gram negative bacteria) belongs
to GHF-42.
Fruits can express multiple β-gal genes. There are
approximately 7 β-gal genes expressed in tomato fruit
development that have amino acid similarity between 33%
and 79%.

11. Sources of β-galactosidase

12. Structure
The 1,023 amino acids of E. coli β-galactosidase were sequenced in
1983, and its determined structure eleven years later in 1994.
The protein is a 464-kDa homotetramer.
Each unit of β-galactosidase consists of five domains.
• Domain 1
is a jelly-roll type β-barrel

13. • Domain 2 and 4
are fibronectin type III-like barrels
• Domain 5
a novel β-sandwich
• Domain 3
is a distorted TIM-type barrel, lacking the fifth helix with a distortion in
the sixth strand. The third domain contains the active site.

14. Identical structure of β-galactosidase
Evolved beta-galactosidase (EBG)
Beta-glucosidase
6-phospho-beta-galactosidase
Beta-mannosidase
Lactase-phlorizin hydrolase

15. Domain 1, blue; Domain 2, green; Domain 3, yellow; Domain 4, cyan; Domain 5, red.
Lighter and darker shades of a given color are used to distinguish the same domain in
different subunits.

16. Mechanism of reaction
It can go through reaction called transgalactosylation
to make allolactose.
Allolactose can also be break to form
monosaccharides. It can also hydrolyze lactose
into galactose and glucose which will go
into glycolysis.

18. Functions
Hydrolyzes the β-glycosidic bond formed between a galactose and its
organic moiety.
In E. coli, the lacZ gene is the structural gene for β-galactosidase;
which is present as part of the inducible system lac operon.
β-galactosidase is vital for organisms as it is a key provider in the
production of energy and a source of carbons through the break
down of lactose to galactose and glucose.
It is also important for the lactose intolerant community as it is
responsible for making lactose-free milk and other dairy products.

19. Applications in food industry

20. Biotechnological applications of β-
galactosidase