What is the Significance of Protein Expression?

The method by which living organisms synthesize proteins and further modify and regulate them is called protein expression. Protein expression plays a significant role in several types of research and is highly utilized in molecular biology, biochemistry, and protein research laboratories.          

Protein Expression  

The central dogma in molecular biology explains the concept of conversion of a double-stranded DNA (deoxyribonucleic acid) to a single-stranded mRNA (messenger ribonucleic acid) and further into a protein. The DNA must be first transcribed to produce mRNA, and then the mRNA must be translated to synthesize proteins. These proteins are unstable and hence have to undergo certain modifications after translation through post-translational modification. Acetylation, folding of proteins, glycosylation, and disulfide bridge formation are some of the post-translational modifications that help a protein in its primary structure attain a stable state and subsequently get converted into a functional state. Thus, the process which synthesizes proteins and modifies them for their conversion into functional proteins is included in the protein expression.   

Investigations on Protein Expression  

Various investigations have been performed and experimented with controlling the expression of proteins for various applications in medicine and biotechnology. The significant investigation and the findings on protein expression are as follows:          

• The in vivo visualization of proteins is identified by the researchers with the help of certain fluorescent proteins. Then, these fluorescent proteins are tagged with the targeted proteins for studying their localization, structure, functions, and interactions.            
• Further, these investigations also favor protein purification, which can be utilized for various researches in molecular biology and the medical field.          
• The proteins can be synthesized easily compared to DNA, which can be achieved with the help of live cells or from the complex mixtures obtained from the cells.          
• The expression system plays a very important role in the production of recombinant proteins and their purification. The investigation has led to the discovery of various systems for expression, including insect, bacterial, mammalian, yeast, cell-free, and plant expression systems capable of synthesizing recombinant proteins.          

Methodology and Factors   

 Methodology steps 

The general methodology behind the expression systems is listed as follows:          

• Cells need to be transfected with the desired DNA template.          
• These cells are allowed for the processes such as transcription, translation, and post-translational modifications.          
• The cells are lysed to extract the protein from them. Protein extraction is facilitated with the help of protein tags.          

The extracted proteins which have several contaminants must be separated from the impurities by the purification methodologies.

In this way, protein is expressed and extracted from a cell for various investigations. The resulted protein is called recombinant protein as its synthesis is invitro.          

Factors         

Among the various systems for expression, deciding a particular system for a specific protein depends on certain factors. The desired protein that has to be expressed, the quantity required of that particular protein, and the downstream technique plans are the significant factors to be considered for choosing a system for expression accordingly. 

Expression Systems      

The expression systems utilized for expressing protein perform their role with the help of plasmids that act as expression vectors. Some of these systems are cell-free, insect, yeast, plant, mammalian, and bacterial expression systems.          

1. Mammalian expression system: This system is preferred to express mammalian proteins, which need several post-translational modifications for perfect functioning. The mammalian expression system synthesizes proteins either through cell lines that are stable or in a transient method. The yield of the protein is higher in both cases if the transfection is achieved correctly. Therefore, it is significant in producing antibodies, functional assays, expressing complex proteins, analyzing structures, studying protein interactions, and producing viruses. Examples of the mammalian expression system are Expi293 and CHO (Chinese Hamster Ovarian Cells).                                        

2. Insect expression system: The eukaryotic protein complex in nature can be expressed in this system with the proper post-translational modifications. The expression system in insects is categorized into two categories such as non-lytic insect cell and baculovirus-infected cell. However, this system is capable of large-scale protein production. Still, it has certain drawbacks, such as the complex cultural conditions similar to the expression system of mammals and the time taken to complete the process. Hi5 cells and Sf21 cells are few examples of the insect expression system.  

3. Plant expression system: It is considered one of the low-tech and cheap systems for the large production of proteins. Cells obtained from plants such as rice, maize, sugarcane, and tobacco are used for expressing proteins. This system shows similar features to the mammalian system. Still, it can be expensive with its purification and extraction techniques and takes a long time because of its biochemical complexity. Nevertheless, the plant expression system is highly recommended for expressing proteins such as pharmaceuticals and antibodies. Interleukins are the specific proteins that can be expressed efficiently with this system.     

4. Bacterial expression system: It is a popular expression system as bacteria are easily culturable and exhibit faster growth. Further, the bacterial expression system is beneficial in producing higher yields. The main drawback of this system is that it cannot perform post-translational modifications, and thus it cannot synthesize functional mammalian proteins that are multi-domain. In addition, the bacteria in certain cases synthesize insoluble proteins, leading to the formation of inclusion bodies. The protein from these inclusion bodies can be extracted only with harsh chemical reagents. Escherichia coliis the popular bacterial system of protein expression. E. coli have been used in various pharmaceutical industries for expressing proteins. pET, pBAD, and pRSET are few popular expression vectors of DNA used in bacterial cells to produce proteins.   

4. Yeast expression system: Many yeast species are used to synthesize recombinant proteins in large quantities. Saccharomyces cerevisiae is the most used system due to its application as a model organism in biochemistry and genetic engineering. However, the growth rate of yeast seems to be very slow, and further, its growth conditions have to be optimized in an often manner.          

5. Cell-free expression system: It is an in-vitro method where the components of translation and transcription are arranged in a purified manner. Ribosomes, tRNAs (transfer RNAs), RNA polymerase, amino acids, and ribonucleotides are the arranged components in the cell-free system for protein synthesis. This system is expensive and has high risk and challenging factors in the technical aspect.

Context and Applications

 This topic is significant in the professional exams for both Bachelors and Master courses related to biology. Some of the courses are listed below: 

• Bachelors of Science in Genetics        
• Bachelors of Science in Biochemistry and Molecular Biology        
• Bachelors of Science in Ecology and Evolutionary Biology        
• Master of Science in Biological Science        
• Master of Science in Biotechnology       
• Master of Science in Proteomics