Barnase (a

portmanteau A portmanteau word, or portmanteau (, ) is a blend of wordsprotein 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 ...

that consists of 110

amino acid Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although hundreds of amino acids exist in nature, by far the most important are the alpha-amino acids, which comprise proteins. Only 22 alpha am ...

s and has

ribonuclease Ribonuclease (commonly abbreviated RNase) is a type of nuclease that catalyzes the degradation of RNA into smaller components. Ribonucleases can be divided into endoribonucleases and exoribonucleases, and comprise several sub-classes within the ...

activity. It is synthesized and secreted by the bacterium ''

Bacillus amyloliquefaciens ''Bacillus amyloliquefaciens'' is a species of bacterium in the genus ''Bacillus'' that is the source of the BamHI restriction enzyme. It also synthesizes a natural antibiotic protein barnase, a widely studied ribonuclease that forms a famously t ...

'', but is lethal to the cell when expressed without its inhibitor

barstar Barstar is a small protein synthesized by the bacterium ''Bacillus amyloliquefaciens''. Its function is to inhibit the ribonuclease activity of its binding partner barnase Barnase (a portmanteau of "BActerial" "RiboNucleASE") is a bacterial pr ...

. The inhibitor binds to and occludes the ribonuclease active site, preventing barnase from damaging the cell's

RNA Ribonucleic acid (RNA) is a polymeric molecule essential in various biological roles in coding, decoding, regulation and expression of genes. RNA and deoxyribonucleic acid ( DNA) are nucleic acids. Along with lipids, proteins, and carbohydra ...

after it has been synthesized but before it has been secreted. The barnase/barstar complex is noted for its extraordinarily tight protein-protein binding, with an

on-rate In chemistry, biochemistry, and pharmacology, a dissociation constant (K_D) is a specific type of equilibrium constant that measures the propensity of a larger object to separate (dissociate) reversibly into smaller components, as when a Complex (c ...

of 10⁸s⁻¹M⁻¹.

Protein folding studies

Barnase has no

disulfide bonds In biochemistry, a disulfide (or disulphide in British English) refers to a functional group with the structure . The linkage is also called an SS-bond or sometimes a disulfide bridge and is usually derived by the coupling of two thiol groups. In ...

, nor does it require

divalent cations In chemistry, the valence (US spelling) or valency (British spelling) of an element is the measure of its combining capacity with other atoms when it forms chemical compounds or molecules. Description The combining capacity, or affinity of an ...

or non-peptide components to fold. This simplicity, in combination with its reversible folding transition, means that barnase has been extensively studied in order to understand how proteins fold. The folding of barnase has been extensively studied in the laboratory of

Alan Fersht Sir Alan Roy Fersht (born 21 April 1943) is a British chemist at the MRC Laboratory of Molecular Biology, Cambridge, and an Emeritus Professor in the Department of Chemistry at the University of Cambridge. He was Master of Gonville and Caius C ...

, who used it as the test case in developing a method of characterizing

protein folding Protein folding is the physical process by which a protein chain is translated to its native three-dimensional structure, typically a "folded" conformation by which the protein becomes biologically functional. Via an expeditious and reproduci ...

transition state In chemistry, the transition state of a chemical reaction is a particular configuration along the reaction coordinate. It is defined as the state corresponding to the highest potential energy along this reaction coordinate. It is often marked wi ...

s known as

phi value analysis Phi value analysis, \phi analysis, or \phi-value analysis is an experimental protein engineering technique for studying the structure of the folding transition state of small protein domains that fold in a two-state manner. The structure of the f ...

Active site and catalytic mechanism

Barnase catalyzes

hydrolysis Hydrolysis (; ) is any chemical reaction in which a molecule of water breaks one or more chemical bonds. The term is used broadly for substitution reaction, substitution, elimination reaction, elimination, and solvation reactions in which water ...

at diribonucleotide GpN sites. Cleavage occurs in two steps using a general acid-base mechanism: a cyclic intermediate is formed during the first

transesterification In organic chemistry, transesterification is the process of exchanging the organic group R″ of an ester with the organic group R' of an alcohol. These reactions are often catalyzed by the addition of an acid or base catalyst. The reaction can ...

step, which is then hydrolysed to release the cleaved

. The two most important residues involved in catalysis are Glu73 and His102, which are both essential for enzymatic activity. Glu73 is the general base whilst His102 is the general acid. Although it is not directly involved in acid-base catalysis, Lys27 is also critical for activity; it has been implicated in transition-state substrate binding.

References

External links

InterPro entry
Ribonucleases Bacterial enzymes {{enzyme-stub

Protein folding studies

Active site and catalytic mechanism

See also

References

Further reading

External links