Chemical modification of Enzymes,Enzyme PEG Conjugates,L asparaginase,Conjugates,Peptide Ligases,Stereoselectivity,Subtilisin

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Home >> Industrial and Microbial Biotechnology >> Protein and Enzymes Engineering >>Chemical modification of Enzymes

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Chemical modification of enzymes
We know that the proteins synthesized under the control of gene sequences in a cell undergo post-translational modification. This leads to stability, structural integrity, altered solubility and viscosity of individual proteins. This may also alter the chemical reactivity. These alterations can be achieved in vitro and may sometimes even create an entirely new enzyme, by creating new active sites or modifying the old ones. Some of the examples will be described in this section.

Enzyme-PEG conjugates.
An enzyme L-asparaginase (isolated from microbes) has anti-tumour properties, but is toxic with a life time of less then 18hr thus reducing its utility. It was shown that E. coli L-asparaginase can be modified by polyethylene glycol derivatives to produce PEG-asparaginase conjugates, which differ from the native enzyme in following features: (i) it retains only 52% of the catalytic activity of native enzyme; (ii) it becomes resistant to proteolytic degradation; (iii) it does not cause allergy. In view of this, PEG-asparaginase has been used to treat malignant murine (mouse), canine (cats, etc.) and human tumours. PEG conjugates of a large number of enzymes (adenosine deaminase, uricase, catalase, etc.) have been prepared and will be utilized in industry also.

L Asparaginase

PEG Asparaginase

Modification of proteases into peptide ligases.
Peptide ligation to native enzymes may lead to high specificity and stereoselectivity, and may suppress side reactions. Therefore, synthesis of any enzyme that may catalyze peptide ligation will be most welcome. Protease 'subtilisin' has been modified (by converting a serine into cysteine or seleno-cysteine) into thiol- and seleno-subtilisin, the two semi-synthetic enzymes (they are damaged proteases), which can catalyse peptide ligation. Both these damaged proteases are efficient peptide ligases. Similarly, histidine residue can also be modified to yield peptide ligases.

Chemical modification of a protease

a protease Subtilisin Intermediate Protein Peptide Ligase

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A two step procedure for producing a nucelase from a DNA binding protein

DNA Binding Protein

1. Addition of a thiol group	2. NH_2n
3. DNA binding protein	4. 1,10 pheneathroline copper complex (a metal chelating agent)

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Coulping of an Oligonucleotide to a Nonspecific Phosphodiesterase to Generate a Site Specific Nuclease

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