RNase A: Difference between revisions

=='''Structure, Catalysis, and Substrate Binding'''==

=='''Structure'''==  

RNase A is made up of a single polypeptide chain of 124 residues. Of the 20 natural amino acids, RNase A possesses 19 of them, excluding tryptophan.<ref name="Raines" /> This single polypeptide chain is cross-linked internally by four disulfide linkages, which contribute to the stability of RNase A. Long four-stranded anti-parallel <scene name='Sandbox_Reserved_192/Beta_sheet/4'>ß-sheets</scene> and three short <scene name='Sandbox_Reserved_192/Alpha_helices/2'>α-helices</scene> make up the <scene name='Sandbox_Reserved_192/Secondary_structure/3'>secondary structure</scene> of RNase A.<ref name="Raines" /> The amino acid sequence was discovered to determine the three-dimensional structure of RNase A by Christian Anfinsen in the 1950s. Urea was used to denature RNase A, and mercaptoethanol was used to reduce and cleave the four disulfide bonds in RNase A to yield eight Cys residues. Catalytic activity was lost due to denaturation. When the urea and mercaptoethanol were removed, the denatured ribonuclease refolded spontaneously into its correct tertiary structure with restoration of its catalytic activity. Disulfide bonds were also reformed in the same position. The Anfinsen experiment provided evidence that the amino acid sequence contained all the information required for the protein to fold into its native three-dimensional structure. Anfinsen received the 1972 Nobel Prize in Chemistry for his work with RNase A. Nevertheless, ensuing work showed some proteins require further assistance, such as molecular chaperones, to fold into their native structure.<ref name="Lehninger" />

=='''Inhibitors'''==

[[Image:RI.PNG|300px|left|thumb|Figure III: Ribonuclease Inhibitor-RNase A Complex. Left, Ribonuclease Inhibitor (RI)is composed of alternating alpha helix (blue) and beta sheets (green). Right, RI-RNase A inhibition forms when RI complex with the active site cleft of RNase (yellow).  Figure generated via ''Pymol'']]

Due to the high rate of RNA hydrolysis by RNase A, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site of RNase A due to its <scene name='User:R._Jeremy_Johnson/RNaseA/Ri_simple/1'>non-globular nature</scene>. RI is a 50 kD protein that is composed of 16 repeating subunits of alpha helices and beta sheets, giving it a noticeable <scene name='User:R._Jeremy_Johnson/RNaseA/Ri_nonglobular/1'>horseshoe like appearance</scene>. The RI-RNase protein-protein interaction has the highest known affinity of any protein-protein interactions with an approximate dissociation constant (''K''d) of 5.8 X 10-14 for almost all types of ribonucleases.<ref>PMID:7877692</ref> The ability to be selective for almost all types of RNases, and yet retain such a high Kd is product of its mechanism of inhibition. The interior residues of the horseshoe shaped RI are able to bind to the charged residues of the active site cleft of RNase A, such as <scene name='User:R._Jeremy_Johnson/RNaseA/Ri_rnasea_lys7_gln11_lys41/1'>Lys7, Gln11, and Lys41 </scene>. By studying the amphibian RNase, Onconase, the residues Lys7 and Gln11 of RNase A were shown to be the most important in this interaction. In onconase, these residues are replaced with non-charged amino acids, which help prevent the binding of RI to the protein <ref>PMID:18930025</ref>

* [http://www.proteopedia.org/wiki/index.php/RNaseS_RNaseB RNase S and RNase B]

* [http://www.proteopedia.org/wiki/index.php/RNaseA_Nobel_Prizes RNase A Nobel Prizes]

== '''References''' ==