Proteomics - Challenges in Delivery of Therapeutic Genomics and Proteomics - page 410

chain groups may not be that critical, because the outcome of the reaction is bound
to be stereospecific. However, lability of enzymes to acids and thermal stresses limit
their utility to a great extent.
Quite a voluminous amount of work has been published on harnessing the
enzymes for peptide and protein synthesis. However, some of the major constraints
associated with enzyme-based peptide synthesis are (1) the need for optimiza-
tion of every peptide bond-forming reaction with the given enzyme under question;
(2) purity of enzymes; and (3) pH, concentrations, and solubilities of products, and
so on. These constraints act as major barriers and elude the commercialization and
scale up of enzymatic methods for protein synthesis. In spite of limitations for the
use of enzymes in peptide synthesis, they may prove to be prospective in the semi-
synthesis of peptides.
In the last decade of the twentieth century, certainly one of the landmark discover-
ies made in human history is drug designing based on rare recombinant proteins for
human disease therapy. These new chemical entities (NCEs) are based on rare human
proteins produced by genetic engineering or recombinant DNA technology. Human
insulin and human growth hormone are some of the revolutionary products that have
been the marvelous gifts of recombinant DNA technology. Looking at the technical
problems associated with delivery of native proteins, recombinant DNA technology
provides a beacon of hope. By applying the basic principles of genetic engineering
and biotechnology, we can generate recombinant proteins to cater to the current and
future needs of diagnostics and therapeutics.
8.6.4 Recombinant Proteins in Heterologous Host/Vector Systems
In the majority of cases, cloning complementary DNA is a major obstacle. To manip-
ulate the faulty protein, we have to replace the gene present in the host vector sys-
tem. A variety of host cell expression vector systems, including prokaryotic systems
of E. coli and eukaryotic systems like yeast ( Saccharomyces cerevisae ) have been
extensively investigated. One of the primary issues in expression vector design is
to provide for replication of vector and gene so that dividing cells will receive the
appropriate genetic information. The gene has to be made devoid of noncoding and
biologically insignificant nucleotide sequences. Sometimes, a case may arise, where
the protein cannot undergo the required post-translational synthesis in bacterial cell
due to lack of favorable conditions. For instance, tissue plasminogen activator (TPA)
normally forms 17 specific intramolecular disulfide bridges. However, only a small
fragment exhibited biological activity when expressed in E. coli due to improper
disulfide bridge formation.
The primary goal of using recombinant DNA technology is to optimize prediction
of natural or native proteins by manipulating the genes. The concept has been suc-
cessfully applied to human insulin, TPA, and growth hormone; however, much needs
to be done in this field. For example, TPA is useful in coronary artery disease and
acute stages of myocardial infarction, however, few complications exist even today.
In spite of biotechnological advancements, issues related to delivery systems, stabil-
ity, specificity, scale up, and so on, remain elusive.