Crossover junction endodeoxyribonuclease, also known as Holliday junction resolvase, Holliday junction endonuclease, Holliday junction-cleaving endonuclease, Holliday junction-resolving endoribonuclease, crossover junction endoribonuclease, and cruciform-cutting endonuclease, is an

enzyme Enzymes () are proteins that act as biological catalysts by accelerating chemical reactions. The molecules upon which enzymes may act are called substrates, and the enzyme converts the substrates into different molecules known as products. A ...

involved in

DNA repair DNA repair is a collection of processes by which a cell identifies and corrects damage to the DNA molecules that encode its genome. In human cells, both normal metabolic activities and environmental factors such as radiation can cause DNA dam ...

and

homologous recombination Homologous recombination is a type of genetic recombination in which genetic information is exchanged between two similar or identical molecules of double-stranded or single-stranded nucleic acids (usually DNA as in cellular organisms but may ...

. Specifically, it performs endonucleolytic cleavage that results in single-stranded crossover between two homologous DNA molecules at the

Holliday junction A Holliday junction is a branched nucleic acid structure that contains four double-stranded arms joined. These arms may adopt one of several conformations depending on buffer salt concentrations and the sequence of nucleobases closest to the ju ...

to produce recombinant DNA products for chromosomal segregation. This process is known as Holliday junction resolution.

Biological Function

The

is a structure that forms during

genetic recombination Genetic recombination (also known as genetic reshuffling) is the exchange of genetic material between different organisms which leads to production of offspring with combinations of traits that differ from those found in either parent. In eukaryo ...

, and links two double-stranded DNA molecules with a single-stranded crossover, which form during

mitotic In cell biology, mitosis () is a part of the cell cycle in which replicated chromosomes are separated into two new nuclei. Cell division by mitosis gives rise to genetically identical cells in which the total number of chromosomes is maintai ...

and

meiotic Meiosis (; , since it is a reductional division) is a special type of cell division of germ cells in sexually-reproducing organisms that produces the gametes, such as sperm or egg cells. It involves two rounds of division that ultimately res ...

recombination. Crossover junction endodeoxyribonucleases catalyze Holiday junction resolution, which is the formation of separate recombinant DNA molecules and chromosomal separation after the crossover event at the Holliday junction. Crossover junction endodeoxyribonucleases with Holliday Junction resolution function have been identified in all three domains of life -

bacteria Bacteria (; singular: bacterium) are ubiquitous, mostly free-living organisms often consisting of one biological cell. They constitute a large domain of prokaryotic microorganisms. Typically a few micrometres in length, bacteria were among ...

archaea Archaea ( ; singular archaeon ) is a domain of single-celled organisms. These microorganisms lack cell nuclei and are therefore prokaryotes. Archaea were initially classified as bacteria, receiving the name archaebacteria (in the Archaebac ...

, and

eukarya Eukaryotes () are organisms whose cells have a nucleus. All animals, plants, fungi, and many unicellular organisms, are Eukaryotes. They belong to the group of organisms Eukaryota or Eukarya, which is one of the three domains of life. Bacte ...

. RuvC in bacteria, CCE1 in

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

, and GEN1 in humans are all crossover junction endodeoxyribonucleases that perform Holliday Junction resolution. Holliday junction resolution catalyzed by crossover junction endodeoxyribonuclease is shown in the figure below. Resolvase function

Crossover junction endodeoxyribonucleases also play key roles in

. During

cell growth Cell growth refers to an increase in the total mass of a cell, including both cytoplasmic, nuclear and organelle volume. Cell growth occurs when the overall rate of cellular biosynthesis (production of biomolecules or anabolism) is greater than ...

and

meiosis Meiosis (; , since it is a reductional division) is a special type of cell division of germ cells in sexually-reproducing organisms that produces the gametes, such as sperm or egg cells. It involves two rounds of division that ultimately resu ...

, DNA double-strand breaks (DSBs) often occur, and are usually repaired by homologous recombination. Because Crossover junction endodeoxyribonucleases perform Holliday Junction resolution, a crucial step of homologous recombination, they are therefore involved in repair of DSBs.

Structure

E. coli ''Escherichia coli'' (),Wells, J. C. (2000) Longman Pronunciation Dictionary. Harlow ngland Pearson Education Ltd. also known as ''E. coli'' (), is a Gram-negative, facultative anaerobic, rod-shaped, coliform bacterium of the genus ''Escher ...

RuvC, a Crossover junction endodeoxyribonuclease, is a small

protein Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, respo ...

of about 20 kD, and its active form is a

dimer Dimer may refer to: * Dimer (chemistry), a chemical structure formed from two similar sub-units ** Protein dimer, a protein quaternary structure ** d-dimer * Dimer model, an item in statistical mechanics, based on ''domino tiling'' * Julius Dimer ...

that requires and binds a

magnesium Magnesium is a chemical element with the symbol Mg and atomic number 12. It is a shiny gray metal having a low density, low melting point and high chemical reactivity. Like the other alkaline earth metals (group 2 of the periodic ta ...

ion RuvC is a 3-layer alpha-beta sandwich with a

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

between 5

alpha-helices The alpha helix (α-helix) is a common motif in the secondary structure of proteins and is a right hand-helix conformation in which every backbone N−H group hydrogen bonds to the backbone C=O group of the amino acid located four residues ear ...

. The enzyme contains two binding channels that contact the backbones of the Holliday junction over seven

nucleotides Nucleotides are organic molecules consisting of a nucleoside and a phosphate. They serve as monomeric units of the nucleic acid polymers – deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), both of which are essential biomolecules w ...

. A Holliday junction resolvase enzyme has also been identified in archaea in

Pyrococcus furiosus ''Pyrococcus furiosus'' is a heterotrophic, strictly anaerobic, extremophilic, model species of archaea. It is classified as a hyperthermophile because it thrives best under extremely high temperatures, and is notable for having an optimum growt ...

cells - it is encoded by a gene called hjc and is composed of 123 amino acids . A figure of

Thermus thermophilus ''Thermus thermophilus'' is a Gram-negative bacterium used in a range of biotechnological applications, including as a model organism for genetic manipulation, structural genomics, and systems biology. The bacterium is extremely thermophilic, w ...

RuvC in complex with a Holliday junction is shown below. 4LD0

Mechanism

These enzymes are highly selective for branched DNA, although

induced fit Enzyme catalysis is the increase in the rate of a process by a biological molecule, an "enzyme". Most enzymes are proteins, and most such processes are chemical reactions. Within the enzyme, generally catalysis occurs at a localized site, calle ...

occurs in the enzyme-substrate (resolvase-Holloday Junction) complex formation. Much remains unknown about the exact mechanism of action, but it is known that bacteria,

bacteriophages A bacteriophage (), also known informally as a ''phage'' (), is a duplodnaviria virus that infects and replicates within bacteria and archaea. The term was derived from "bacteria" and the Greek φαγεῖν ('), meaning "to devour". Bacterio ...

and archaea catalyze Holliday junction resolution by introducing symmetric nicks across the Holliday junction . Analysis of crossover junction endodeoxyribonucleases from bacteriophages (T7 endonuclease I), bacteria (RuvC),

fungi A fungus ( : fungi or funguses) is any member of the group of eukaryotic organisms that includes microorganisms such as yeasts and molds, as well as the more familiar mushrooms. These organisms are classified as a kingdom, separately from ...

(GEN1) and humans (hMus81-Eme1) have revealed that the enzymes function in dimers, and part of the resolution reaction takes place in a partially dissociated enzyme-substrate intermediate.

Human Relevance

After a 20-year search, in 2008, a human crossover junction endodeoxyribonuclease, GEN1, was finally identified . GEN1 performs similar functions and operates by similar mechanisms as previously studied Crossover junction endodeoxyribonuclease in bacteria, archaea, and other eukarya. The enzyme is thought to play a role in

Bloom's syndrome Bloom syndrome (often abbreviated as BS in literature) is a rare autosome, autosomal recessive genetic disorder characterized by short stature, predisposition to the development of cancer, and genomic instability. BS is caused by mutations in the ' ...

. It has been proposed that Bloom's syndrome involves the induction of DSBs via an unidentified Holliday junction resolvase. It has also been shown that overexpression of Holliday Junction resolvase function is correlated with RAD51-overexpressing

cancers Cancer is a group of diseases involving Cell growth#Disorders, abnormal cell growth with the potential to Invasion (cancer), invade or Metastasis, spread to other parts of the body. These contrast with benign tumors, which do not spread. Poss ...

References

External links

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