Lactase

Template:Short description Template:Cs1 config Template:Distinguish Template:Infobox enzyme Template:Infobox protein

Lactase (Template:EnzExplorer) is an enzyme produced by many organisms and is essential to the complete digestion of whole milk. It breaks down the sugar lactose into its component parts, galactose and glucose. Lactase is found in the brush border of the small intestine of humans and other mammals. People deficient in lactase or lacking functional lactase may experience the symptoms of lactose intolerance after consuming milk products.<ref name="pmid19639477">Template:Cite journal</ref> Microbial β-galactosidase (often loosely referred to as lactase) can be purchased as a food supplement and is added to milk to produce "lactose-free" milk products.

UsesEdit

Food useEdit

Lactase is an enzyme that some people are unable to produce in their small intestine.<ref>{{#invoke:citation/CS1|citation |CitationClass=web }}</ref> Technology to produce lactose-free milk, ice cream, and yogurt was developed by the USDA Agricultural Research Service in 1985.<ref>{{#invoke:citation/CS1|citation |CitationClass=web }}</ref>Lactase is added to milk, thereby hydrolyzing the lactose in the milk, leaving it slightly sweet but digestible by everyone.<ref>Template:Cite news</ref> Without lactase, lactose-intolerant people pass the lactose undigested to the colon<ref>{{#invoke:citation/CS1|citation |CitationClass=web }}</ref> where bacteria break it down, creating carbon dioxide which leads to bloating and flatulence.

Medical useEdit

Lactase supplements can be used to treat lactose intolerance.<ref name=NIH2014Dig>{{#invoke:citation/CS1|citation |CitationClass=web }}</ref>

Industrial useEdit

Lactase produced commercially can be extracted both from yeasts such as Kluyveromyces fragilis and Kluyveromyces lactis and from molds, such as Aspergillus niger and Aspergillus oryzae.<ref>Template:Cite journal</ref> Its primary commercial use in supplements is to break down lactose in milk to make it suitable for people with lactose intolerance.<ref>{{#invoke:citation/CS1|citation |CitationClass=web }}</ref><ref>Template:Cite book</ref> The U.S. Food and Drug Administration has not independently evaluated these products.<ref>{{#invoke:citation/CS1|citation |CitationClass=web }}</ref>

Lactase (or a similar form of β-galactosidase) is also used to screen for blue white colonies in the multiple cloning sites of various plasmid vectors in Escherichia coli or other bacteria.<ref>{{#invoke:citation/CS1|citation |CitationClass=web }}</ref><ref>{{#invoke:citation/CS1|citation |CitationClass=web }}</ref>

MechanismEdit

The temperature optimum for human lactase is about 37 °C<ref name="pmid17512743">Template:Cite journal</ref> and the pH optimum is 6.<ref name="pmid6786877">Template:Cite journal</ref>

In metabolism, the β-glycosidic bond in D-lactose is hydrolyzed to form D-galactose and D-glucose, which can be absorbed through the intestinal walls and into the bloodstream. The overall reaction that lactase catalyzes is as follows:

C₁₂H₂₂O₁₁ + H₂O → C₆H₁₂O₆ + C₆H₁₂O₆ + heat.

lactose + H₂O → β-D-galactose + D-glucose

The catalytic mechanism of D-lactose hydrolysis retains the substrate anomeric configuration in the products.<ref name = "doi10.1021/bi011727i">Template:Cite journal</ref> While the details of the mechanism are uncertain, the stereochemical retention is achieved via a double displacement reaction. Studies of E. coli lactase have proposed that hydrolysis is initiated when a glutamate nucleophile on the enzyme attacks from the axial side of the galactosyl carbon in the β-glycosidic bond.<ref name = "pmid11732897">Template:Cite journal</ref> The removal of the D-glucose leaving group may be facilitated by Mg-dependent acid catalysis.<ref name = "pmid11732897"/> The enzyme is liberated from the α-galactosyl moiety upon equatorial nucleophilic attack by water, which produces D-galactose.<ref name = "doi10.1021/bi011727i"/>

Substrate modification studies have demonstrated that the 3′-OH and 2′-OH moieties on the galactopyranose ring are essential for enzymatic recognition and hydrolysis.<ref name = "pmid7648581">Template:Cite journal</ref> The 3′-hydroxy group is involved in initial binding to the substrate while the 2′- group is not necessary for recognition but needed in subsequent steps. This is demonstrated by the fact that a 2-deoxy analog is an effective competitive inhibitor (K_i = 10mM).<ref name = "pmid7648581"/> Elimination of specific hydroxyl groups on the glucopyranose moiety does not eliminate catalysis.<ref name = "pmid7648581"/>

Proposed mechanism of lactose hydrolysis by Lactase enzyme

Lactase also catalyzes the conversion of phlorizin to phloretin and glucose.

Commercial lactase is used as a medication for lactose intolerance. Since it is an enzyme, its function can be inhibited by the acidity of the stomach. However, it is packaged in an acid-proof tablet, allowing the enzyme to pass through the stomach intact and remain in the small intestine. In the small intestine it can act on ingested lactose molecules, allowing the body to absorb the digested sugar which would otherwise cause cramping and diarrhea. Since the enzyme is not absorbed, it is excreted.Template:Cn

Structure and biosynthesisEdit

Preprolactase, the primary translation product, has a single polypeptide primary structure consisting of 1927 amino acids.<ref name="pmid2460343">Template:Cite journal</ref> It can be divided into five domains: (i) a 19-amino-acid cleaved signal sequence; (ii) a large prosequence domain that is not present in mature lactase; (iii) the mature lactase segment; (iv) a membrane-spanning hydrophobic anchor; and (v) a short hydrophilic carboxyl terminus.<ref name = "pmid2460343"/> The signal sequence is cleaved in the endoplasmic reticulum, and the resulting 215-kDa pro-LPH is sent to the Golgi apparatus, where it is heavily glycosylated and proteolytically processed to its mature form.<ref>Template:Cite journal</ref> The prodomain has been shown to act as an intramolecular chaperone in the ER, preventing trypsin cleavage and allowing LPH to adopt the necessary 3-D structure to be transported to the Golgi apparatus.<ref>Template:Cite journal</ref>

Schematic of processing and localization of human lactase translational product

Mature human lactase consists of a single 160-kDa polypeptide chain that localizes to the brush border membrane of intestinal epithelial cells. It is oriented with the N-terminus outside the cell and the C-terminus in the cytosol.<ref name = "pmid2460343"/> LPH contains two catalytic glutamic acid sites. In the human enzyme, the lactase activity has been connected to Glu-1749, while Glu-1273 is the site of phlorizin hydrolase function.<ref>Template:Cite journal</ref>

Genetic expression and regulationEdit

In humans, lactase is encoded by a single genetic locus on chromosome 2.<ref name = "pmid9148757">Template:Cite journal</ref> It is expressed exclusively by mammalian small intestine enterocytes and in very low levels in the colon during fetal development.<ref name="pmid9148757"/> Humans are born with high levels of lactase expression. In most of the world's population, lactase transcription is down-regulated after weaning, resulting in diminished lactase expression in the small intestine,<ref name = "pmid9148757"/> which causes the common symptoms of adult-type hypolactasia, or lactose intolerance.<ref>{{#invoke:citation/CS1|citation |CitationClass=web }}</ref> The LCT gene provides the instructions for making lactase. Lactose intolerance in infants (congenital lactase deficiency) is caused by mutations in the LCT gene. Mutations are believed to interfere with the function of lactase, causing affected infants to have a severely impaired ability to digest lactose in breast milk or formula.<ref name="medlineplus.gov">{{#invoke:citation/CS1|citation |CitationClass=web }}</ref>

Some population segments exhibit lactase persistence resulting from a mutation that is postulated to have occurred 5,000–10,000 years ago, coinciding with the rise of cattle domestication.<ref>Template:Cite journal</ref> This mutation has allowed almost half of the world's population to metabolize lactose without symptoms. Studies have linked the occurrence of lactase persistence to two different single-nucleotide polymorphisms about 14 and 22 kilobases upstream of the 5'-end of the LPH gene.<ref>Template:Cite journal</ref> Both mutations, C→T at position -13910 and G→ A at position -22018, have been independently linked to lactase persistence.<ref name = "pmid15777735">Template:Cite journal</ref>

The lactase promoter is 150 base pairs long and is located upstream of the site of transcription initiation.<ref name = "pmid15777735"/> The sequence is highly conserved in mammals, suggesting that critical cis-transcriptional regulators are located nearby.<ref name = "pmid15777735"/> Cdx-2, HNF-1α, and GATA have been identified as transcription factors.<ref name = "pmid15777735"/> Studies of hypolactasia onset have demonstrated that despite polymorphisms, little difference exists in lactase expression in infants, showing that the mutations become increasingly relevant during development.<ref name = "pmid9609760">Template:Cite journal</ref> Developmentally regulated DNA-binding proteins may down-regulate transcription or destabilize mRNA transcripts, causing decreased LPH expression after weaning.<ref name = "pmid9609760"/>