Sandbox Reserved 197
From Proteopedia
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Ribonuclease A is an enzyme involved in catalyzing RNA degradation. The structure of RNase A has been determined through crystallography [http://en.wikipedia.org/wiki/Crystallography] and FABMS [http://en.wikipedia.org/wiki/Fast_atom_bombardment]. RNase A is composed of four anti-parallel B-sheets and 3 a-helixes. The <scene name='Sandbox_Reserved_197/Active_site/1'>active site</scene> lies within the cleft and houses three residues important for catalysis: His12, His119, and Lys41. Presence of eight cysteine residues that form four disulfide bonds and four ''cis'' proline residues greatly effect the structure and folding kinetics of RNase A. | Ribonuclease A is an enzyme involved in catalyzing RNA degradation. The structure of RNase A has been determined through crystallography [http://en.wikipedia.org/wiki/Crystallography] and FABMS [http://en.wikipedia.org/wiki/Fast_atom_bombardment]. RNase A is composed of four anti-parallel B-sheets and 3 a-helixes. The <scene name='Sandbox_Reserved_197/Active_site/1'>active site</scene> lies within the cleft and houses three residues important for catalysis: His12, His119, and Lys41. Presence of eight cysteine residues that form four disulfide bonds and four ''cis'' proline residues greatly effect the structure and folding kinetics of RNase A. | ||
| - | == '''Folding''' == | + | == '''Protein Folding''' == |
| - | + | Interatomic interactions are responsible for formation of a protein's 3D structure [http://en.wikipedia.org/wiki/Protein_folding]. Several of these interactions have been identified by the use of site directed mutagenesis to wildtype RNase A and subsequent comparison of the crystal structure to the wildtype. | |
==='''Proline Conformation'''=== | ==='''Proline Conformation'''=== | ||
| - | + | The presence of ''cis'' [http://en.wikipedia.org/wiki/Cis_configuration]proline residues plays a large role in protein folding. In nature, most amino acids reside in a ''trans'' conformation, but due to their cyclic structure, prolines are more stable in a ''cis'' conformation. RNase A contains four proline residues, two reside in the ''cis'' conformation and two in the ''trans'' conformation. Importance of these conformations are demonstrated with several mutations to the wilde type. | |
| + | The <scene name='Sandbox_Reserved_197/Tyr92-pro93/5'>Tyr92-Pro93</scene> peptide group of RNase A in its native state is found in the ''cis'' conformation. Despite a mutation from proline to alanine, <scene name='Sandbox_Reserved_197/P93a/7'>P93A</scene>, a ''cis'' conformation still forms; this is an unfavorable conformation for an alanine residue. Upon unfolding, Tyr92-Ala93 undergoes isomerization to form its more favorable ''trans'' conformation demonstrating that the ''cis'' conformation is favored by protein interactions other than the proline residue. Although the overall structure of RNase A is not affected by this mutation, the rate of folding greatly decreases upon insertion of the P93A mutation. | ||
<scene name='Sandbox_Reserved_197/Cis-proline114/1'>''cis'' proline</scene> | <scene name='Sandbox_Reserved_197/Cis-proline114/1'>''cis'' proline</scene> | ||
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==='''Disulfide Bonds'''=== | ==='''Disulfide Bonds'''=== | ||
Another important feature of the folding of RNase A is the presence of four disulfide bonds. These bonds contribute to the thermal stability and the rate of folding of RNase A. The residues involved in these linkages include <scene name='Sandbox_Reserved_197/Cys26-cys84/4'>Cys26-Cys84</scene>, <scene name='Sandbox_Reserved_197/Cys58-cys110/4'>Cys58-Cys110</scene>, <scene name='Sandbox_Reserved_197/40-95_disulfide_native_form/4'>Cys40-Cys95</scene>, and <scene name='Sandbox_Reserved_197/Cys65-cys72/5'>Cys65-Cys72</scene>. Cys26-Cys84 and Cys58-Cys110 create an interaction between an α-helix and a β-sheet. This connection is the main contributor to the thermodynamic stability. RNase A actually has a rate-determining three-disulfide intermediate. An analog of this, <scene name='Sandbox_Reserved_197/C40-95a_variant/6'>C[40,95]A</scene>, shows RNase A, missing the disulfide bond, Cys40-Cys95, that would normally occur here. As you can see in the variant, there are only 3 disulfide bonds present, shown in red. | Another important feature of the folding of RNase A is the presence of four disulfide bonds. These bonds contribute to the thermal stability and the rate of folding of RNase A. The residues involved in these linkages include <scene name='Sandbox_Reserved_197/Cys26-cys84/4'>Cys26-Cys84</scene>, <scene name='Sandbox_Reserved_197/Cys58-cys110/4'>Cys58-Cys110</scene>, <scene name='Sandbox_Reserved_197/40-95_disulfide_native_form/4'>Cys40-Cys95</scene>, and <scene name='Sandbox_Reserved_197/Cys65-cys72/5'>Cys65-Cys72</scene>. Cys26-Cys84 and Cys58-Cys110 create an interaction between an α-helix and a β-sheet. This connection is the main contributor to the thermodynamic stability. RNase A actually has a rate-determining three-disulfide intermediate. An analog of this, <scene name='Sandbox_Reserved_197/C40-95a_variant/6'>C[40,95]A</scene>, shows RNase A, missing the disulfide bond, Cys40-Cys95, that would normally occur here. As you can see in the variant, there are only 3 disulfide bonds present, shown in red. | ||
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| + | ==='''Summary'''=== | ||
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==''Medical Importance''== | ==''Medical Importance''== | ||
Revision as of 12:30, 1 April 2011
| This Sandbox is Reserved from Feb 02, 2011, through Jul 31, 2011 for use by the Biochemistry II class at the Butler University at Indianapolis, IN USA taught by R. Jeremy Johnson. This reservation includes Sandbox Reserved 191 through Sandbox Reserved 200. |
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Contents |
Introduction
Ribonuclease A is an enzyme involved in catalyzing RNA degradation. The structure of RNase A has been determined through crystallography [1] and FABMS [2]. RNase A is composed of four anti-parallel B-sheets and 3 a-helixes. The lies within the cleft and houses three residues important for catalysis: His12, His119, and Lys41. Presence of eight cysteine residues that form four disulfide bonds and four cis proline residues greatly effect the structure and folding kinetics of RNase A.
Protein Folding
Interatomic interactions are responsible for formation of a protein's 3D structure [3]. Several of these interactions have been identified by the use of site directed mutagenesis to wildtype RNase A and subsequent comparison of the crystal structure to the wildtype.
Proline Conformation
The presence of cis [4]proline residues plays a large role in protein folding. In nature, most amino acids reside in a trans conformation, but due to their cyclic structure, prolines are more stable in a cis conformation. RNase A contains four proline residues, two reside in the cis conformation and two in the trans conformation. Importance of these conformations are demonstrated with several mutations to the wilde type. The peptide group of RNase A in its native state is found in the cis conformation. Despite a mutation from proline to alanine, , a cis conformation still forms; this is an unfavorable conformation for an alanine residue. Upon unfolding, Tyr92-Ala93 undergoes isomerization to form its more favorable trans conformation demonstrating that the cis conformation is favored by protein interactions other than the proline residue. Although the overall structure of RNase A is not affected by this mutation, the rate of folding greatly decreases upon insertion of the P93A mutation.
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Disulfide Bonds
Another important feature of the folding of RNase A is the presence of four disulfide bonds. These bonds contribute to the thermal stability and the rate of folding of RNase A. The residues involved in these linkages include , , , and . Cys26-Cys84 and Cys58-Cys110 create an interaction between an α-helix and a β-sheet. This connection is the main contributor to the thermodynamic stability. RNase A actually has a rate-determining three-disulfide intermediate. An analog of this, , shows RNase A, missing the disulfide bond, Cys40-Cys95, that would normally occur here. As you can see in the variant, there are only 3 disulfide bonds present, shown in red.
Summary
Medical Importance
Protein folding, along with its inhibitions, is immensely important to the human. Such diseases as ALS, Alzheimer's Disease, and Parkinson's Disease can all be traced back to the protein. Proteins can form aberrant aggregates when they do not fold correctly. This abnormaility can be fatally toxic to the human nerve cells. During folding, proteins sometimes make a mistake. Each protein contains . The hydrophilic residues lie on the outer part of the protein and the hydrophobic residues bury themselves due to the hydrophobic effect [5]. In the case of these aggregates, the mistake exposes "sticky" of the interior that can cause several proteins to stick to one another. In the future researchers hope to design drugs that combat this mistake in the protein folding. The use of ribonuclease A in protein folding research has been an instrumental feature in designing experiments to determine these "misfolding" snapshots and in developing therapies to prevent this problem in the future.
