β-galactosidase enzyme

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.

3. 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.

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

5. Inhibitors of β- galactosidase Beta-gal is inhibited by L-ribose,  N on-competitive inhibitor iodine, C ompetitive inhibitors  2-phenylethyl 1-thio-beta-D-galactopyranoside (PETG), D- galactonolactone ,  isopropyl thio-beta-D- galactoside (IPTG), galactose

6. 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.

7. 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 

8. Conti …. E. Coli  belongs to GHF-2, all plants belong to GHF-35, and  Thermus thermophilus (gram negative bacteria)  belongs to GHF-42.  F ruits 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 %.

9. Sources of β- galactosidase

10. 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

11. 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. 

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

13. 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.

14. 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 .

15. Functions H ydrolyzes 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 .

16. Applications in food industry

17. Biotechnological applications of β- galactosidase