Sandbox Reserved 198: Difference between revisions
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=='''Structure equals Function'''== | =='''Structure equals Function'''== | ||
The synthesis of semisynthetic RNasa A clearly exhibits the structure to function relationship that defines proteins. In the RNase A protein, the removal of six C terminal residues, leaving <scene name='Sandbox_Reserved_198/Rnase_1-118/1'>RNase 1-118</scene>, completely halts enzymatic activity | The synthesis of semisynthetic RNasa A clearly exhibits the structure to function relationship that defines proteins. In the RNase A protein, the removal of six C terminal residues, leaving <scene name='Sandbox_Reserved_198/Rnase_1-118/1'>RNase 1-118</scene>, completely halts enzymatic activity (Martin, 1987). However, a complex of RNase 1-118 with a synthetic polypeptide comprising the <scene name='Sandbox_Reserved_198/Synthetic_component/3'>C terminal residues, 111-124 Component</scene> restores enzymatic activity to RNase A. Upon the addition of the synthetic chain, the semisynthetic enzyme adopts a structure that closely resembles that of natural RNase (Martin, 1987). The restoration of the structure reconstitutes the enzymatic activity of RNase to 98% (Martin, 1987). | ||
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=='''Synthetic Method'''== | =='''Synthetic Method'''== | ||
The RNase 1-118 was prepared by successive digestion of RNase A pepsin and carboxypeptidase A (Doscher, 1983). The synthetic component, RNase 111-124, was prepared by the use of solid-phase peptide synthetic | Solid-Phase Peptide Synthesis | ||
Peptide synthesis is the production of proteins in which multiple amino acids are linked together through peptide bonds. A general chemical requirement for peptide synthesis is the blockage of the carboxyl group of one amino acid and the amino group of the second amino acid. The carboxyl group of the free carboxyl group can be activated and the new peptide bond is formed (Merrifield, 1984). A common type of peptide synthesis is the solid-phase synthesis, in which the end of the peptide chain is attached to a solid support, as shown in Figure 1. | |||
The semi-synthetic RNase A comprises of residues 1-118 and the synthetic analog of residues 111-124. The RNase 1-118 was prepared by successive digestion of RNase A pepsin and carboxypeptidase A (Doscher, 1983). The synthetic component, RNase 111-124, was prepared by the use of solid-phase peptide synthetic methods, in which the peptide chain was assembled in the stepwise manner while it was attached at one end to a solid support. The peptide chain was extended by repetitive steps of de-protection, neutralization and coupling until the desired sequence was obtained (Lin, 1970). It was important that the synthesis proceeds rapidly and in high yields to prevent side reactions or by-products. | |||
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1. Introduction to Ribonuclease A by Raines: http://www.uta.edu/faculty/sawasthi/Enzymology-4351-5324/Class%20Syllabus%20Enzymology/ribonucleaseA.pdf | 1. Introduction to Ribonuclease A by Raines: http://www.uta.edu/faculty/sawasthi/Enzymology-4351-5324/Class%20Syllabus%20Enzymology/ribonucleaseA.pdf | ||
2. Solid Phase Synthesis by Merrifield (Nobel Prize Winner):http://nobelprize.org/nobel_prizes/chemistry/laureates/1984/merrifield-lecture.pdf | 2. Introduction to Peptide Synthesis: http://en.wikipedia.org/wiki/Solid_phase_peptide_synthesis#Solid-phase_synthesis | ||
3.Solid Phase Synthesis by Merrifield (Nobel Prize Winner):http://nobelprize.org/nobel_prizes/chemistry/laureates/1984/merrifield-lecture.pdf | |||
4. Chemical Synthesis of Proteins:http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2845543/?tool=pmcentrez | |||
5. Refined Crystal Structure: http://www.ncbi.nlm.nih.gov/pubmed/3680234 | |||
'''References''' | '''References''' | ||
Martin, Philip D., Marilynn S. Doscher, and Brian F. P. Edwards. "The Redefined Crystal Structure of a Fully Active Semisynthetic Ribonuclease at 1.8-A Resolution." The Journal of Biological Chemistry 262.33 (1987): 15930-5938. | Martin, Philip D., Marilynn S. Doscher, and Brian F. P. Edwards. "The Redefined Crystal Structure of a Fully Active Semisynthetic Ribonuclease at 1.8-A Resolution." The Journal of Biological Chemistry 262.33 (1987): 15930-5938. | ||
Marilynn S. Doscher, Philip D. Martin and Brian F.P. Edwards, "Characerization of the Histidine Proton Nuclear Magnetic Resonance of a Semisynthetic Ribonuclease." Biochemistry, 1983,22,4125-4131. | Marilynn S. Doscher, Philip D. Martin and Brian F.P. Edwards, "Characerization of the Histidine Proton Nuclear Magnetic Resonance of a Semisynthetic Ribonuclease." Biochemistry, 1983,22,4125-4131. | ||
Merrifield B. "Solid Phase Synthesis", Nobel Lecture, 8 December, 1984. | |||
Lin, M. C. (1970) Journal of Biological Chemistry, 245, 6726-6731. | Lin, M. C. (1970) Journal of Biological Chemistry, 245, 6726-6731. | ||