Galactokinase is an

enzyme An enzyme () is a protein that acts as a biological catalyst by accelerating chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates into different mol ...

(phosphotransferase) that facilitates the

phosphorylation In biochemistry, phosphorylation is described as the "transfer of a phosphate group" from a donor to an acceptor. A common phosphorylating agent (phosphate donor) is ATP and a common family of acceptor are alcohols: : This equation can be writ ...

of α-D-galactose to

galactose 1-phosphate D-Galactose-1-phosphate is an intermediate in the intraconversion of glucose and uridine diphosphate galactose. It is formed from galactose by galactokinase.The improper metabolism of galactose-1-phosphate is a characteristic of galactosemia. The ...

at the expense of one

molecule A molecule is a group of two or more atoms that are held together by Force, attractive forces known as chemical bonds; depending on context, the term may or may not include ions that satisfy this criterion. In quantum physics, organic chemi ...

of ATP. Galactokinase catalyzes the second step of the

Leloir pathway The Leloir pathway is a metabolic pathway for the catabolism of D-galactose. It is named after Luis Federico Leloir, who first described it. In the first step, galactose mutarotase facilitates the conversion of β-D-galactose to α-D-galactose s ...

, a

metabolic pathway In biochemistry, a metabolic pathway is a linked series of chemical reactions occurring within a cell (biology), cell. The reactants, products, and Metabolic intermediate, intermediates of an enzymatic reaction are known as metabolites, which are ...

found in most

organisms An organism is any living thing that functions as an individual. Such a definition raises more problems than it solves, not least because the concept of an individual is also difficult. Many criteria, few of them widely accepted, have been pr ...

for the

catabolism Catabolism () is the set of metabolic pathways that breaks down molecules into smaller units that are either oxidized to release energy or used in other anabolic reactions. Catabolism breaks down large molecules (such as polysaccharides, lipid ...

of α-D-galactose to

glucose 1-phosphate Glucose 1-phosphate (also called Cori ester) is a glucose molecule with a phosphate group on the 1'-carbon. It can exist in either the α- or β- anomeric form. Reactions of α-glucose 1-phosphate Catabolic In glycogenolysis, it is the direct p ...

. First isolated from

mammal A mammal () is a vertebrate animal of the Class (biology), class Mammalia (). Mammals are characterised by the presence of milk-producing mammary glands for feeding their young, a broad neocortex region of the brain, fur or hair, and three ...

ian

liver The liver is a major metabolic organ (anatomy), organ exclusively found in vertebrates, which performs many essential biological Function (biology), functions such as detoxification of the organism, and the Protein biosynthesis, synthesis of var ...

, galactokinase has been studied extensively in

yeast Yeasts are eukaryotic, single-celled microorganisms classified as members of the fungus kingdom (biology), kingdom. The first yeast originated hundreds of millions of years ago, and at least 1,500 species are currently recognized. They are est ...

archaea Archaea ( ) is a Domain (biology), domain of organisms. Traditionally, Archaea only included its Prokaryote, prokaryotic members, but this has since been found to be paraphyletic, as eukaryotes are known to have evolved from archaea. Even thou ...

plants Plants are the eukaryotes that form the kingdom Plantae; they are predominantly photosynthetic. This means that they obtain their energy from sunlight, using chloroplasts derived from endosymbiosis with cyanobacteria to produce sugars f ...

, and

humans Humans (''Homo sapiens'') or modern humans are the most common and widespread species of primate, and the last surviving species of the genus ''Homo''. They are Hominidae, great apes characterized by their Prehistory of nakedness and clothing ...

Structure

Galactokinase is composed of two domains separated by a large cleft. The two regions are known as the N- and

C-terminal The C-terminus (also known as the carboxyl-terminus, carboxy-terminus, C-terminal tail, carboxy tail, C-terminal end, or COOH-terminus) is the end of an amino acid chain (protein or polypeptide), terminated by a free carboxyl group (-COOH). When t ...

domains, and the

adenine Adenine (, ) (nucleoside#List of nucleosides and corresponding nucleobases, symbol A or Ade) is a purine nucleotide base that is found in DNA, RNA, and Adenosine triphosphate, ATP. Usually a white crystalline subtance. The shape of adenine is ...

ring of ATP binds in a hydrophobic pocket located at their interface. The

N-terminal The N-terminus (also known as the amino-terminus, NH2-terminus, N-terminal end or amine-terminus) is the start of a protein or polypeptide, referring to the free amine group (-NH2) located at the end of a polypeptide. Within a peptide, the amin ...

domain is marked by five strands of mixed

beta-sheet The beta sheet (β-sheet, also β-pleated sheet) is a common structural motif, motif of the regular protein secondary structure. Beta sheets consist of beta strands (β-strands) connected laterally by at least two or three backbone chain, backbon ...

and five

alpha-helices An alpha helix (or α-helix) is a sequence of amino acids in a protein that are twisted into a coil (a helix). The alpha helix is the most common structural arrangement in the secondary structure of proteins. It is also the most extreme type of l ...

, and the C-terminal domain is characterized by two layers of anti-parallel beta-sheets and six alpha-helices. Galactokinase does not belong to the

sugar Sugar is the generic name for sweet-tasting, soluble carbohydrates, many of which are used in food. Simple sugars, also called monosaccharides, include glucose Glucose is a sugar with the Chemical formula#Molecular formula, molecul ...

kinase In biochemistry, a kinase () is an enzyme that catalyzes the transfer of phosphate groups from high-energy, phosphate-donating molecules to specific substrates. This process is known as phosphorylation, where the high-energy ATP molecule don ...

family, but rather to a class of ATP-dependent enzymes known as the GHMP superfamily. GHMP is an abbreviation referring to its original members: galactokinase,

homoserine kinase In enzymology, a homoserine kinase () is an enzyme that catalyzes the chemical reaction :ATP + L-homoserine \rightleftharpoons ADP + O-phospho-L-homoserine Thus, the two substrates of this enzyme are ATP and L-homoserine, whereas its two prod ...

mevalonate kinase Mevalonate kinase is an enzyme (specifically a kinase) that in humans is encoded by the ''MVK'' gene In biology, the word gene has two meanings. The Mendelian gene is a basic unit of heredity. The molecular gene is a sequence of nucleotid ...

, and

phosphomevalonate kinase Phosphomevalonate kinase is an enzyme () in the mevalonate Mevalonic acid (MVA) is a key organic compound in biochemistry; the name is a contraction of dihydroxymethylvalerolactone. The carboxylate anion of mevalonic acid, which is the pred ...

. Members of the GHMP superfamily have great three-dimensional similarity despite only ten to 20% sequence identity. These enzymes contain three well-conserved motifs (I, II, and III), the second of which is involved in

nucleotide Nucleotides are Organic compound, organic molecules composed of a nitrogenous base, a pentose sugar and a phosphate. They serve as monomeric units of the nucleic acid polymers – deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), both o ...

binding and has the sequence

Pro Pro is an abbreviation meaning "professional". Pro, PRO or variants thereof might also refer to: People * Miguel Pro (1891–1927), Mexican priest * Pro Hart (1928–2006), Australian painter * Mlungisi Mdluli (born 1980), South African ret ...

-X-X-X- Gly- Leu-X- Ser-Ser- Ala.

Sugar specificity

Galactokinases across different species display a great diversity of substrate specificities. ''

E. coli ''Escherichia coli'' ( )Wells, J. C. (2000) Longman Pronunciation Dictionary. Harlow ngland Pearson Education Ltd. is a gram-negative, facultative anaerobic, rod-shaped, coliform bacterium of the genus ''Escherichia'' that is commonly foun ...

'' galactokinase can also phosphorylate 2-deoxy-D-galactose, 2-amino-deoxy-D-galactose, 3-deoxy-D-galactose and D-fucose. The enzyme cannot tolerate any C-4 modifications, but changes at the C-2 position of D-galactose do not interfere with enzyme function. Both human and

rat Rats are various medium-sized, long-tailed rodents. Species of rats are found throughout the order Rodentia, but stereotypical rats are found in the genus ''Rattus''. Other rat genera include '' Neotoma'' (pack rats), '' Bandicota'' (bandicoo ...

galactokinases are also able to successfully phosphorylate 2-deoxy-D-galactose. Galactokinase from ''

S. cerevisiae ''Saccharomyces cerevisiae'' () (brewer's yeast or baker's yeast) is a species of yeast (single-celled fungal microorganisms). The species has been instrumental in winemaking, baking, and brewing since ancient times. It is believed to have been o ...

'', on the other hand, is highly specific for D-galactose and cannot phosphorylate

glucose Glucose is a sugar with the Chemical formula#Molecular formula, molecular formula , which is often abbreviated as Glc. It is overall the most abundant monosaccharide, a subcategory of carbohydrates. It is mainly made by plants and most algae d ...

mannose Mannose is a sugar with the formula , which sometimes is abbreviated Man. It is one of the monomers of the aldohexose series of carbohydrates. It is a C-2 epimer of glucose. Mannose is important in human metabolism, especially in the glycosylatio ...

arabinose Arabinose is an aldopentose – a monosaccharide containing five carbon atoms, and including an aldehyde (CHO) functional group. Properties For biosynthetic reasons, most saccharides are almost always more abundant in nature as the "D"-form, o ...

, fucose,

lactose Lactose is a disaccharide composed of galactose and glucose and has the molecular formula C12H22O11. Lactose makes up around 2–8% of milk (by mass). The name comes from (Genitive case, gen. ), the Latin word for milk, plus the suffix ''-o ...

galactitol Galactitol (dulcitol) is a sugar alcohol, the reduction product of galactose. It has a slightly sweet taste. In people with galactokinase deficiency, a form of galactosemia, excess dulcitol forms in the lens of the eye leading to cataracts. Gal ...

, or 2-deoxy-D-galactose. Moreover, the kinetic properties of galactokinase also differ across species. The sugar specificity of galactokinases from different sources has been dramatically expanded through

directed evolution Directed evolution (DE) is a method used in protein engineering that mimics the process of natural selection to steer proteins or nucleic acids toward a user-defined goal. It consists of subjecting a gene to iterative rounds of mutagenesis (cre ...

and structure-based

protein engineering Protein engineering is the process of developing useful or valuable proteins through the design and production of unnatural polypeptides, often by altering amino acid sequences found in nature. It is a young discipline, with much research taking pl ...

. The corresponding broadly permissive sugar anomeric kinases serve as a cornerstone for ''in vitro'' and ''in vivo''

glycorandomization Glycorandomization, is a drug discovery and drug development technology platform to enable the rapid diversification of bioactive small molecules, drug leads and/or approved drugs through the attachment of sugars. Initially developed as a facile me ...

Mechanism

Recently, the roles of

active site In biology and biochemistry, the active site is the region of an enzyme where substrate molecules bind and undergo a chemical reaction. The active site consists of amino acid residues that form temporary bonds with the substrate, the ''binding s ...

residues in human galactokinase have become understood. Asp-186 abstracts a

proton A proton is a stable subatomic particle, symbol , Hydron (chemistry), H+, or 1H+ with a positive electric charge of +1 ''e'' (elementary charge). Its mass is slightly less than the mass of a neutron and approximately times the mass of an e ...

from C1-OH of α-D-galactose, and the resulting

alkoxide In chemistry, an alkoxide is the conjugate base of an alcohol and therefore consists of an organic group bonded to a negatively charged oxygen atom. They are written as , where R is the organyl substituent. Alkoxides are strong bases and, whe ...

nucleophile In chemistry, a nucleophile is a chemical species that forms bonds by donating an electron pair. All molecules and ions with a free pair of electrons or at least one pi bond can act as nucleophiles. Because nucleophiles donate electrons, they are ...

attacks the γ-

phosphorus Phosphorus is a chemical element; it has Chemical symbol, symbol P and atomic number 15. All elemental forms of phosphorus are highly Reactivity (chemistry), reactive and are therefore never found in nature. They can nevertheless be prepared ar ...

of ATP. A

phosphate group Phosphates are the naturally occurring form of the element phosphorus. In chemistry, a phosphate is an anion, salt, functional group or ester derived from a phosphoric acid. It most commonly means orthophosphate, a derivative of orthophosp ...

is transferred to the sugar, and Asp-186 may be

deprotonated Deprotonation (or dehydronation) is the removal (transfer) of a proton (or hydron, or hydrogen cation), (H+) from a Brønsted–Lowry acid in an acid–base reaction.Henry Jakubowski, Biochemistry Online Chapter 2A3, https://employees.csbsju.ed ...

water Water is an inorganic compound with the chemical formula . It is a transparent, tasteless, odorless, and Color of water, nearly colorless chemical substance. It is the main constituent of Earth's hydrosphere and the fluids of all known liv ...

. Nearby Arg-37 stabilizes Asp-186 in its

anion An ion () is an atom or molecule with a net electrical charge. The charge of an electron is considered to be negative by convention and this charge is equal and opposite to the charge of a proton, which is considered to be positive by conven ...

ic form and has also been proven to be essential to galactokinase function in

point mutation A point mutation is a genetic mutation where a single nucleotide base is changed, inserted or deleted from a DNA or RNA sequence of an organism's genome. Point mutations have a variety of effects on the downstream protein product—consequences ...

experiments. Both the aspartic acid and arginine active site residues are highly conserved among galactokinases. Galactokinase crystal structure

Biological function

The Leloir pathway catalyzes the conversion of galactose to glucose. Galactose is found in

dairy products Dairy products or milk products are food products made from (or containing) milk. The most common dairy animals are cow, water buffalo, goat, nanny goat, and Sheep, ewe. Dairy products include common grocery store food around the world such as y ...

, as well as in

fruits In botany, a fruit is the seed-bearing structure in flowering plants (angiosperms) that is formed from the ovary after flowering. Fruits are the means by which angiosperms disseminate their seeds. Edible fruits in particular have long propaga ...

and

vegetables Vegetables are edible parts of plants that are consumed by humans or other animals as food. This original meaning is still commonly used, and is applied to plants collectively to refer to all edible plant matter, including flowers, fruits, ...

, and can be produced endogenously in the breakdown of

glycoproteins Glycoproteins are proteins which contain oligosaccharide (sugar) chains covalently attached to amino acid side-chains. The carbohydrate is attached to the protein in a cotranslational or posttranslational modification. This process is known a ...

and

glycolipids Glycolipids () are lipids with a carbohydrate attached by a glycosidic (covalent) bond. Their role is to maintain the stability of the cell membrane and to facilitate cellular recognition, which is crucial to the immune response and in the co ...

. Three enzymes are required in the Leloir pathway: galactokinase,

galactose-1-phosphate uridylyltransferase Galactose-1-phosphate uridyltransferase (or GALT, G1PUT) is an enzyme () responsible for converting ingested galactose to glucose. Galactose-1-phosphate uridyltransferase (GALT) catalyzes the second step of the Leloir pathway of galactose meta ...

, and UDP-galactose 4-epimerase. Galactokinase catalyzes the first committed step of galactose catabolism, forming galactose 1-phosphate.

Disease relevance

Galactosemia Galactosemia (British galactosaemia, from Greek γαλακτόζη + αίμα, meaning galactose + blood, accumulation of galactose in blood) is a rare genetics, genetic Metabolism, metabolic Disease, disorder that affects an individual's ability t ...

, a rare

metabolic disorder A metabolic disorder is a disorder that negatively alters the body's processing and distribution of macronutrients, such as proteins, fats, and carbohydrates. Metabolic disorders can happen when abnormal chemical reactions in the body alter the ...

characterized by decreased ability to metabolize galactose, can be caused by a mutation in any of the three enzymes in the Leloir pathway.

Galactokinase deficiency Galactokinase deficiency is an autosome, autosomal dominance (genetics), recessive inborn errors of metabolism, metabolic disorder marked by an accumulation of galactose and galactitol secondary to the decreased conversion of galactose to galactos ...

, also known as galactosemia type II, is a

recessive In genetics, dominance is the phenomenon of one variant (allele) of a gene on a chromosome masking or overriding the effect of a different variant of the same gene on the other copy of the chromosome. The first variant is termed dominant and ...

metabolic disorder caused by a

mutation In biology, a mutation is an alteration in the nucleic acid sequence of the genome of an organism, virus, or extrachromosomal DNA. Viral genomes contain either DNA or RNA. Mutations result from errors during DNA or viral replication, ...

in human galactokinase. About 20 mutations have been identified that cause galactosemia type II, the main

symptom Signs and symptoms are diagnostic indications of an illness, injury, or condition. Signs are objective and externally observable; symptoms are a person's reported subjective experiences. A sign for example may be a higher or lower temperature ...

of which is early onset

cataracts A cataract is a cloudy area in the lens of the eye that leads to a decrease in vision of the eye. Cataracts often develop slowly and can affect one or both eyes. Symptoms may include faded colours, blurry or double vision, halos around ligh ...

. In

lens A lens is a transmissive optical device that focuses or disperses a light beam by means of refraction. A simple lens consists of a single piece of transparent material, while a compound lens consists of several simple lenses (''elements'') ...

cells Cell most often refers to: * Cell (biology), the functional basic unit of life * Cellphone, a phone connected to a cellular network * Clandestine cell, a penetration-resistant form of a secret or outlawed organization * Electrochemical cell, a d ...

of the human

eye An eye is a sensory organ that allows an organism to perceive visual information. It detects light and converts it into electro-chemical impulses in neurons (neurones). It is part of an organism's visual system. In higher organisms, the ey ...

aldose reductase In enzymology, aldose reductase (or aldehyde reductase) () is an enzyme in humans encoded by the gene AKR1B1. It is an cytosolic NADPH-dependent oxidoreductase that catalyzes the reduction of a variety of aldehydes and carbonyls, including monos ...

converts galactose to

. As galactose is not being catabolized to glucose due to a galactokinase mutation, galactitol accumulates. This galactitol gradient across the lens cell membrane triggers the

osmotic Osmosis (, ) is the spontaneous net movement or diffusion of solvent molecules through a selectively-permeable membrane from a region of high water potential (region of lower solute concentration) to a region of low water potential (region o ...

uptake of water, and the swelling and eventual

apoptosis Apoptosis (from ) is a form of programmed cell death that occurs in multicellular organisms and in some eukaryotic, single-celled microorganisms such as yeast. Biochemistry, Biochemical events lead to characteristic cell changes (Morphology (biol ...

of lens cells ensues.

References

External links

* {{Portal bar, Biology, border=no EC 2.7.1 Moonlighting proteins