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CHAPTER – I INTRODUCTIION AND IMPORTANCE OF BIOCHEMISTRY Biochemistry, as the name implies, is the chemistry of living organisms. It bridges the gap between the conventional chemistry and biology. organisms have certain extraordinary properties. Living They can grow, respond to stimuli and replicate themselves with high fidelity. All these activities are ultimately interpretable in chemical terms. The lifeless organic molecules with appropriate complexity and properties make a living thing. The basic phenomena of biochemistry are to understand how the collections of inanimate molecules that constitute living organisms interact with each other to maintain life. The basic life processes or chemistry remains broadly the same whether it is a unicellular microorganism or the higher organisms such as human or plants. Life is nothing but thousands of ordered chemical reactions. In other words, chemistry is the logic of all biological phenomena. Biochemistry deals with the chemical nature and chemical behaviour of the living organisms. The word biochemistry is derived from bios – meaning life. The metabolic processes are studied in this branch. Today, there are three main types of biochemistry. Plant biochemistry involves the study of the biochemistry of autotrophic organisms such as photosynthesis and other plant specific biochemical processes. General biochemistry encompasses both plant and animal biochemistry. Human/medical/medicinal biochemistry focuses on the biochemistry of humans and medical illnesses. There is tremendous importance of biochemistry in agriculture. The chemical processes leading to metabolism of protein, carbohydrate, oils, vitamins and minerals, apart from secondary products from plant such as gum, mucilage, oleoresins, terpenes, essential oils, phenolics, alkaloids etc. are more important for farmers and end users for quality of crop and end products of the plants. The genetic engineering and biotechnology involve a great deal of biochemistry for crop improvement. 1.1 History of biochemistry Only during 17th and 18th centuries, important foundations were laid in many fields of biology. The 19th century observed the development of very crucial concepts, which include the cell theory by Schleiden and Schwann, Mendel’s study of inheritance and Darwin’s theory of evolution. The real push to biochemistry was given in 1828 when total synthesis of urea from lead cyanate and ammonia was successfully achieved by Wohler who thus initiated the synthesis of organic compound from inorganic compound. Louis Pasteur, during 1857, did a great deal of work on fermentations and pointed out categorically the central importance of enzymes in this process. The breakthrough in enzyme research and hence, biochemistry was made in 1897 by Edward Buckner when he extracted enzyme from yeast cells in crude form which could ferment a sugar molecule into alcohol. Neuberg introduced the term biochemistry in 1903. The early part of 20th century witnessed a sudden outburst of knowledge in chemical analysis, separation methods, electronic instrumentation for biological studies (X-ray diffraction, electron microscope, etc) which ultimately resulted in understanding the structure and function of several key molecules involved in life processes such as proteins, enzymes, DNA and RNA. In 1926, James Sumner established the protein nature of enzyme. He was responsible for the isolation and crystallization of urease, which provided a breakthrough in studies of the properties of specific enzymes. The first metabolic pathway elucidated was the glycolytic pathway during the first half of the 20th century by Embden and Meyerhof. Otto Warburg, Cori and Parnas also made very important contributions relating to glycolytic pathway. Krebs established the citric acid and urea cycles during 1930-40. In 1940, Lipmann described the central role of ATP in biological systems. The biochemistry of nucleic acids entered into a phase of exponential growth after the establishment of the structure of DNA in 1953 by Watson and Crick followed by the discovery of DNA polymerase by Kornberg in 1956. From 1960 onwards, biochemistry plunged into an interdisciplinary phase sharing much in common with biology and molecular genetics. Frederick Sanger’s contributions in the sequencing of protein in 1953 and nucleic acid in 1977 were responsible for further developments in the field of protein and nucleic acid research. 2 The growth of biochemistry and molecular biology was phenomenal during the past two decades. The development of recombinant DNA research by Snell and coworkers during 1980 allowed for further growth and emergence of a new field, the genetic engineering. Thus there was progressive evolution of biology to biochemistry and then to molecular biology, genetic engineering and biotechnology. The major contributions made by some of the Scientists in the developments of biochemistry are listed below in table 1.1. Table 1.1 Scientists and their major contribution to biochemistry and related fields. 1780-1789 Lavoisier 1828 Wohler 1837 Berzelius 1838 1854-1864 Schleiden and Schwann Louis Pasteur 1866 Mendel 1869 1877 1894 Miescher Kuhne Emil Fischer 1897 Buckner 1902 1903 1905 Emil Fischer Neuberg Harden and Young 1912 1913 Neuberg Michaelis and Menten Sumner 1926 1933 Embden Meyerhof Recognized that respiration is oxidation and first measured oxygen consumption by human subject Synthesized the first organic compound, urea from inorganic components Postulated the catalytic nature of fermentation. He also identified lactic acid as a product of muscle activity. Enunciated the cell theory Proved that fermentation is caused by microorganisms Reported the principles of segregation and independent assortment of genes Discovered DNA Proposed the term ‘Enzyme’ Demonstrated the specificity of enzymes and the lock and key relationship between enzyme and substrate Discovered alcoholic fermentation in cell-free yeast extract Demonstrated that proteins are polypeptides First used the term ‘biochemistry’ Showed the requirement of phosphate in alcoholic fermentation and identified first coenzyme, cozymase, later shown to be NAD Proposed chemical pathway for fermentation Developed kinetic theory of enzyme action First crystallized an enzyme, urease and proved it to be a protein Demonstrated crucial intermediates in the chemical and pathway of glycolysis and fermentation 3 1937 1940 1940 1944 1948 1950 1950-1953 1953 1953 1954 Parnas Krebs Lipmann Beadle and Tatum Avery, MacLeod and McCarty Calvin and Benson Pauling and Corey Chargaff Sanger and Thompson Watson and Crick Arnon and Colleagues Discovered citric acid cycle Role of ATP in biological systems Deduced one gene-one enzyme relationship Demonstrated that bacterial transformation was caused by DNA Discovered that phosphoglyceric acid is an early intermediate in photosynthetic CO2 fixation Proposed the -helix structure for keratins Discovered the base composition of DNA Determined the complete amino acid sequence of insulin Proposed the double-helical model for DNA structure Discovered photosynthetic phosphorylation 1956 Kornberg Discovered DNA polymerase 1958 Meselson and Stahl Hamilton and Daniel Nathans Jacob & Monod Confirmed the Watson-Crick conservative replication of DNA Restriction endonucleases 1961 Nirenberg and Matthaei 1961-1965 Nirenberg Khorana and Ochoa Reported that polyuridylic acid codes for phenylalanine and this opened the way to identification of genetic code Identified the genetic code words for amino acids 1969 Arber Restriction endonucleases 1977 Sanger Determination of DNA sequence 1980 Snell Development of recombinant DNA research leading to genetic engineering 1984 Kary Mullis Polymerase chain reaction 1997 Wilmut 1999 Ingo potrykus Viable offspring derived from fetal and adult mammalian cells. Golden rice rich in -carotene 2006 Andrew Z. Fire Nobel Prize for discovering the role of RNA and Craig C. interference (RNAi), in the silencing of gene Mello expression 1960 1961 model of semi Proposed the operon hypothesis and postulated the function of messenger RNA 4 In 20th century, due to development of analytical instruments, rapid progress in biochemistry was made. The use of Radio Isotope labelling studies / Mass Spectroscopy / Electrophoresis / Spectrophotometry / NMR and genetic engineering methods speeded up the research on biochemistry. We are inan exciting time in Plant Biochemistry. The next century is anticipated to be "The Century of Biology" with advances in biotechnology expected to lead economic development. Not only will we be able use advances in Plant Biochemistry to produce better, more nutritious foods which will improve human health more than any other medical advance, but we are on the verge of using plants as sophisticated chemical factories to produce all manner of materials useful to medicine and industry. We are beginning to understand the structure and function of an increasing number of enzymes and other proteins and this is spawning the development of rational design of enzyme function and other methods of protein engineering. A growing number of prokaryote and lower eukaryote genomes have already been completely sequenced and a near complete sequence of the Arabidopsis genome was published in December, 2000. The entire human genome DNA completely sequenced and published in March 2003. We are moving from structural to functional genomics with this latter field being particular relevant to biochemistry. "Information about the function of a protein is contained not in its primary sequence but in its structure, so the classical 'protein folding problem' - how proteins reach their native, folded state from the unfolded, newly synthesized polypeptide - now takes on new significance. Double bond is introduced into fatty acids in plants by a soluble stromal ACP desaturase. This desaturase normally has high specificity for 18:0-ACP and 9. inserts a cis A small number of plant species in several families posses other acyl-ACP desaturases. Coriandrum sativum 4-palmitoyl (16:0)-ACP desaturase that inserts a double bond at carbon 4 of 16:0. Thunbergia alata (black-eyed 5 6-16:0-ACP 9-myristoyl desaturase and Pelargonium xhortorum (14:0)-ACP desaturase. By comparing the protein sequences of these 3 soluble desaturases and R. communis 9-18:0-ACP desaturase and analyzing 3-dimensional structural information from the crystal structure of the R. communis 9-18:0-ACP desaturase, Cahoon et al. were able to decipher the molecular basis for chainlength recognition and positional placement of double bonds into fatty acids. They showed that the regio-specificities could be modified by replacement of specific amino acid residues and that acyl-ACP activities can be rationally redesigned. 9-18:0-ACP desaturase contains a catalytic diiron cluster (and all the other above-mentioned soluble desaturase contain the conserved iron-binding domains) that represents a fixed point for double bond introduction. Adjacent to these iron atoms is a deep, narrow channel that Cahoon et al. predicted was the binding pocket for the 18:0 portion of the substrate. The channel forces the 18:0 to bend at carbons 9-10 that corresponds to the cis configuration of the 18:1 9 product. Thus the architecture of the substrate-binding channel appeared to determine substrate acyl-chain length, position and stereochemistry of the introduced double bond. 9-18:0-ACP desaturase substrate channel with the 18:0 substrate docked is shown in Fig. 3 of Cahoon et al. They predicted that substitution of alanine-188 for the smaller glycine (G188) and tyrosine- 6-16:0-ACP desaturase would extend the cavity at the bottom of the desaturase active site enough to permit 2 more carbon atoms at the methyl end of 18:0-ACP. They in fact found that mutant A188G/Y189F is able to desaturate 18:0-ACP at a similar rate as with 16:0-ACP. 1. It is useful in the treatment of animals and man. 2. It is helpful to fix the need of food and nutrients in animals, so that the malnutrition in population may be avoided. 3. It is helpful in developing the strategy for higher production of crops. 4. Used in science of pharmacology and drugs. 5. Preservation of fruits and vegetables can be prolonged with the help of biochemistry knowledge. 6 6. Its study is helpful in understanding chemical changes during storage and processing of food material. 7. The plant breeding and genetics branch of agriculture is based on biochemical mechanism of genes. 8. Nutrition and care of cattle, cows and buffaloes 9. Metabolisms of different food nutrients are studied with the help of biochemistry. 10. The problems of biology and medical science can be solved. 11. Nutritional deficiency diseases, malnutrition, planning of balanced diet and energy requirement of human being involve the knowledge of biochemistry. 12. Enzymatic reaction of metabolic pathway and its regulation for higher productivity i.e. photosynthesis, reduction of respiration rate. 13. To improve the quality of starch through modification of regulatory enzyme 14. To assess the nutritional quality of starch, protein and carbohydrates etc of products. 15. To enhance the N fixation in plants 7 Biochemistry Introduction Overview and Introduction to Biochemistry By Anne Marie Helmenstine, Ph.D. Biochemistry is the study of the molecules of life, such as DNA. Ben Mills See More About biochemistry biochemist organic chemistry chemistry disciplines Biochemistry is the science in which chemistry is applied to the study of living organisms and the atoms and molecules which comprise living organisms. Take a closer look at what biochemistry is and why the science is important. What Is Biochemistry? Biochemistry is the study of the chemistry of living things. This includes organic molecules and their chemical reactions. Most people consider biochemistry to be synonymous with molecular biology. What Types of Molecules Do Biochemists Study? The principal types of biological molecules, or biomolecules are: carbohydrates lipids proteins nucleic acids Many of these molecules are complex molecules called polymers, which are made up of monomer subunits. Biochemical molecules are based on carbon. What Is Biochemistry Used For? Biochemistry is used to learn about the biological processes which take place in cells and organisms. Biochemistry may be used to study the properties of biological molecules, for a variety of purposes. For example, a biochemist may study the characteristics of the keratin in hair so that a shampoo may be developed that enhances curliness or softness. Biochemists find uses for biomolecules. For example, a biochemist may use a certain lipid as a food additive. 8 Alternatively, a biochemist might find a substitute for a usual biomolecule. For example, biochemists help to develop artificial sweeteners. Biochemists can help cells to produce new products. Gene therapy is within the realm of biochemistry. The development of biological machinery falls within the realm of biochemistry. What Does a Biochemist Do? Many biochemists work in chemistry labs. Some biochemists may focus on modeling, which would lead them to work with computers. Some biochemists work in the field, studying a biochemical system in an organism. Biochemists typically are associated with other scientists and engineers. Some biochemists are associated with universities and they may teach in addition to conducting research. Usually their research allows them to have a normal work schedule, based in one location, with a good salary and benefits. What Disciplines Are Related to Biochemistry? Biochemistry is closely related to other biological sciences that deal with molecules. There is considerable overlap between these disciplines: Molecular Genetics Pharmacology Molecular Biology Chemical Biology When you major in agricultural biochemistry you explore the unknown, the unseen, and the undiscovered wonders of the natural world. Your coursework will provide a foundation in chemistry, physics, mathematics, and biology, as those fields relate to agricultural and biological sciences. Biochemists study plant, animal, and microbial metabolism as well as the structure and biological function of nucleic acids, proteins, carbohydrates, and lipids by using modern techniques such as x-ray crystallography and NMR spectroscopy. Biochemistry is fundamental to modern biotechnology. As an agricultural biochemistry student, you stand on the frontier of scientific discoveries that change our understanding of the world: new approaches to diabetes, nutrition for athletes, developments in genetically engineered, insect-resistant plants, and methods for detecting vitamin and mineral deficiencies. Most agricultural biochemistry graduates continue their training to pursue careers in agricultural and biological sciences and in human and veterinary medicine. 9 10 11 12 13 14 15 16