Write an essay in up to 2000 words on the following topics, describing the procedures that can be used to isolate proteins from a cell and the methods involved in checking the purity. Using diagrams to help, you should explain fully the principles behind each technique. ‘Healthcare Associated Infections ‘ The Procedures are rapidly increasing in today’s world as technology enhances isolating protein from Cells is getting easier and much more precise.
All cells have a structure and a function; all living organisms have cells with unique structures and functions. There are 2 types of cells, these are prokaryotic and eukaryotic cells, both of them are enveloped by a membrane which regulates entry and exit of substances through the cell as well as separating the interior to the external environment. Each cell is made up of different components such as organelles such as mitochondria, nucleus, golgi apparatus, rough and smooth endoplasmic reticulum. Cells also contain proteins. The behaviour of the cells is down to the information coded in genes. The complex structures of proteins allow them to have a variety of functions throughout the cell, they act as enzymes and use their binding ability to attach to specific substrates, proteins are important as they can oligmerise to form fibrils and some are integrated into the membrane which act as protein channels, receptors and cell signalling.
In Healthcare the main procedures used are: 1. Percipation 2. Chromatography 3. centrifugation In healthcare cells are very important as they carry important information about the human body, as most disease is discovered in the blood as cells are infected. The isolation of proteins from cells is very important as we are able to examine and treat infections, viruses etc. with the growing technology in healthcare and treatment becoming widely available the isolation techniques vary.
Infections associated with health care are a growing factor as they can live in the body undetected for a long time, however the detection of these infections is vital so that we can treat them successfully, the cells contain different proteins therefore the correct method is necessary. The problem we face with these infections is to keep the premises clean and free from germs which lead to the infections. Patients who have a long stay at the hospital tend to be victims to these infections as they are around them more often. One of the major infections is the MRSA, (Methicillin-resistant Staphylococcus aureus) this tends to effect:
In science Protein purification involves a number of processes that aim to isolate the desired protein from a complex mixture. It is necessary in order to be able to characterise the function, structure and interactions of the protein of interest, Biological or microbial cultures are usually the starting material for the process. Cell lysis is the initial step in which the protein is freed from the matrix in which it is confined as the cell membrane is disrupted and its internal contents released into a solution known as crude lysate. Ultracentrifugation is used to purify this mixture which then separates the protein from other components including non-protein constituents of the mixture and finally isolated from other proteins that are present. Fractionates of the various cellular components are produced, fractions include soluble proteins, membrane proteins, nucleic acids and cellular organelles.
The proteins from the lysate can be concentrated using precipitation. There are many Different types of chromatography for protein isolation. Protein separation methods make use of difference in molecular weight, chemical properties (such as net charge) and binding affinity of proteins. Different types of gel electrophoresis is utilised (molecular weight and isoelectric point of protein must be known.) to monitor the level of purification and to check the purity. If the protein has distinguishable sprectroscopy features, spectroscopy can be used to monitor level of purification. various enzyme assays used if protein has enzymatic activity. Electrofocusing can be used to isolate proteins according to their charge, all methods are proven and are currently used in labs to separate and examine proteins from each other.
As there are many proteins, they must be extracted from the starting material, which are usually cells; an efficient protocol needs to be developed in order to release the protein by cell disruption. Cell lysis techniques are widely used, this method involves repeated freezing and thawing of tissue or cells containing required the protein. Homogenization is used for large cells broken by chopping action; this is carried out on muscle tissue as well as plant and animal tissue. Hand homogenization is common as it is where cells are forced through narrow gaps which result in the disruption of cell membrane, for example the liver tissue.