Proteins

Protein is a macronutrient that is essential to building muscle mass. Macronutrients provide calories or energy. There are three macronutrients i.e., proteins, carbohydrates and fat. Proteins are large biomolecules or macromolecules that are comprised of one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalyzing metabolic reactions, DNA replication, responding to stimuli, providing structure to cells and organisms, and transporting molecules from one location to another. Cells generally contain thousands of different proteins, each with a different biological activity. A linear chain of amino acid residues is called a polypeptide. Proteins can be very long polypeptide chains of 100 to several thousand amino acid residues. Short polypeptides, containing less than 20–30 residues, are commonly called peptides, or sometimes oligopeptides. There are generally four recognized levels of protein structure i.e., primary structure, secondary structure, tertiary structure and quaternary structure. The primary structure consists of a sequence of amino acids linked together by peptide bonds and includes any disulphide bonds. Most proteins fold into unique 3D structures. The shape into which a protein naturally folds is known as its native conformation. Although many proteins can fold unassisted, simply through the chemical properties of their amino acids, others require the aid of molecular chaperones to fold into their native states.

Protein purification is a series of processes intended to isolate one or a few proteins from a complex mixture, usually cells, tissues or whole organisms. The purification process may separate the protein and non-protein parts of the mixture, and finally separate the desired protein from all other proteins. A wide range of chromatographic procedures include ion-exchange chromatography, size -exclusion chromatography, affinity-chromatography and high performance liquid chromatography make use of differences in size, binding affinities, charge and other properties. Proteins are separated on the basis of mass and charge by electrophoresis. The principle of electrophoresis relies on the movement of a charged ion in an electric field. In practice, the proteins are denatured in a solution containing a detergent (SDS). The proteins in SDS-PAGE are separated on the sole basis of their size. Purification of proteins can be monitored by assaying specific activity.

The function of a protein depends on its amino acid sequence. Differences in protein function result from differences in amino acid composition and sequence. To sequence an entire polypeptide, a chemical method devised by Pehr Edman is usually employed. Short proteins and peptides can be chemically synthesized.

The three dimensional structure of a protein is determined by its amino acid sequence. The spatial arrangement of atoms in a protein is called its conformation. Proteins in any of their functional, folded conformations are called native proteins. A protein's conformation is stabilized by weak interactions. Secondary structure of proteins refers to regular, recurring arrangements in space of adjacent amino acid residues in a polypeptide chain. The most common secondary structures are the alpha-heilx, the beta conformation and beta turns. The protein's tertiary structure shows the overall three-dimensional arrangement of all atoms in a protein. The quaternary structure of a protein is the association of several protein chains or subunits into a closely packed arrangement. It includes organizations from simple dimers to large homo-oligomers and complexes with defined or variable numbers of subunits. The subunits are held together by hydrogen bonds and van der Waals forces between nonpolar side chains.

Biosynthesis of Lipids

Lipids are the principal form of stored energy in most organisms and major constituents of cellular membranes. The ability to synthesize a variety of lipids is essential to all organisms. Fatty acid synthesis is the creation of fatty acids from acetyl-CoA and NADPH through the action of enzymes called fatty acid synthases. This process takes place in the cytoplasm of the cell. Fatty acid biosynthesis requires the participation of a three carbon intermediate, malonyl-CoA, which is not involved in fatty acid breakdown. The synthesis of malonyl-CoA from acetyl-CoA is catalyzed by the enzyme acetyl-CoA carboxylase in an irreversible process. The long carbon chains of fatty acids are assembled in a repeating four step sequence. Each malonyl group and acetyl group is activated by a thioester that links it to fatty acid synthase (multi enzyme complex). Condensation of an acetyl group from acetyl-CoA (activated acyl group) and two carbon atoms derived from malonyl-CoA, with elimination of carbon dioxide from the malonyl group, extends the acyl chain by two carbons. When the chain length reaches 16 carbons, the product leaves the cycle. The four-step process of fatty acid synthesis is the same in all organisms.

Most of the fatty acids synthesized by an organism either get incorporated into triacylglycerols for the storage of metabolic energy or get incorporated into the phospholipid components of membranes. Animals can synthesize and store large quantities of triacylglycerols, to be used later as fuel. Triacylglycerols are formed by the reaction of two molecules of fatty acyl-CoA with glycerol 3-phosphate to form phosphatidic acid, which is dephosphorylated to a diacylglycerol and then acylated by a third molecule of fatty-acyl CoA to yield a triacylglycerol. The rate of triacylglycerol biosynthesis is altered by the action of several hormones.

In eukaryotic cells, phospholipid synthesis occurs primarily on the surfaces of smooth endoplasmic reticulum and the mitochondrial inner membrane. The principal precursors of glycerophospholipids are diacylglycerols. Membrane lipids are insoluble in water, so they can't simply diffuse from the endoplasmic reticulum to their point of insertion. So they are delivered in membrane vesicles that bud from the Golgi complex then move to and fuse with the target membrane.

Cholesterol is an essential molecule in many animals, including humans but is not required in the mammalian diet. Cholesterol plays an important role as a component of cellular membranes and as a precursor of steroid hormones and bile acids. Biosynthesis of cholesterol generally takes place in the endoplasmic reticulum of hepatic cells and begins with acetyl- CoA, which is mainly derived from an oxidation reaction in the mitochondria. Cholesterol is essential for all animal life, with each cell capable of synthesizing it by way of a complex 37-step process. This begins with the mevalonate or HMG-CoA reductase pathway, the target of statin drugs, which encompasses the first 18 steps. This is followed by 19 additional steps to convert the resulting lanosterol into cholesterol. The steroid hormones glucocorticoids, mineralcorticoids and sex hormones are produced from cholesterol by alteration of the side chain and introduction of oxygen atoms into the steroid ring system.

Biosynthesis of amino acids and nucleotides

Amino acids and nucleotides are charged molecules, so their levels must be regulated to maintain electrochemical balance in the cell. The biosynthetic pathways for amino acids and nucleotides share a requirement for nitrogen. The nitrogen cycle maintains a pool of biologically available nitrogen. Air is the most important source of nitrogen but conversion of atmospheric nitrogen into useful forms for living organisms is done by relatively few species. Atmospheric nitrogen is fixed by nitrogen-fixing bacteria to produce ammonia is the first step in the nitrogen cycle. Soil bacteria oxidize ammonia to nitrite (NO2-) and nitrite to nitrate (NO3-) and the process is known as nitrification. Plants and many bacteria can take up and readily reduce nitrate and nitrite through the action of nitrate and nitrite reductases. The ammonia so formed is incorporated into amino acids by plants. Soil bacteria maintain a balance between fixed nitrogen and atmospheric nitrogen by converting nitrate to N2 (nitrogen), the process is called dentrification. Biological nitrogen fixation is carried out by a highly conserved complex of proteins called the nitrogenase complex. In living systems, reduced nitrogen is incorporated first into amino acids and then into other biomolecules, including nucleotides. Glutamate and glutamine are the nitrogen donors, in different biosynthetic reactions. Glutamine synthase catalyzes the synthesis of glutamine from glutamate.

Plants and bacteria synthesize all twenty common amino acids. Out of the twenty basic amino acids, humans are unable to synthesize eight amino acids. Amino acids that must be obtained from the diet are called essential amino acids. Nonessential amino acids are produced in the body, and can be synthesized in simple pathways In addition, the amino acids arginine, cysteine, glycine, glutamine, histidine, proline, serine, and tyrosine are considered conditionally essential, meaning they are not normally required in the diet but must be supplied exogenously to specific populations that do not synthesize it in adequate amounts. The regulation of the synthesis of glutamate from α-ketoglutarate is subject to regulatory control of the Citric Acid Cycle as well as mass action dependent on the concentrations of reactants involved due to the reversible nature of the transamination and glutamate dehydrogenase reactions. The conversion of glutamate to glutamine is regulated by glutamine synthetase (GS) and is a key step in nitrogen metabolism. The amino acid biosynthetic pathways are subject to allosteric end-product inhibition; the regulatory enzyme is usually the first in the sequence.

Many important biomolecules are derived from amino acids. Glycine is a precursor of porphyrins. Glutathione is an important cellular reducing agent formed from three amino acids. Many plant substances are produced by aromatic amino acids. Nitric oxide, a biological messenger is produced from arginine.

Nucleotides are the precursors of nucleic acids (DNA and RNA). They are essential carriers of chemical energy. There are two pathways for nucleotide biosynthesis i.e., de-novo pathways and salvage pathways. The de-novo pathways are nearly identical in all living organisms. The purine ring system is built up step by step beginning with 5-phosphoribosylamine. The amino acids glutamine, glycine and aspartate furnish all the nitrogen atoms of purines. Pyrimidines are synthesized from carbamoyl phosphate and aspartate, and the attachment of ribose-5-phosphate yields the pyrimidine ribonucleotides. Uric acid and urea are the end products of purine and pyrimidine degradation.

Hormones and their regulation

An essential characteristic of multicellular organisms is cell differentiation and division of labor. The specialized functions of the tissues and organs of complex organisms impose characteristic fuel requirements and patterns of metabolism. Hormonal signals integrate and coordinate the metabolic activities of different tissues and optimize the allocation of fuels and precursors to each organ. Hormones are chemical messengers that are secreted directly into the blood, which carries them to organs and tissues of the body to exert their functions. Hormones are required for the correct development of both animals and plants. Hormones affect distant cells by binding to specific receptor proteins in the target cell, resulting in a change in cell function. When a hormone binds to the receptor, it results in the activation of a signal transduction pathway that typically activates gene transcription, resulting in increased expression of target proteins. Mammals have several classes of hormones, distinguishable by their chemical structures and their modes of action. Some hormones such as peptide, amine, and eicosanoid hormones act from outside the target cells through surface receptors and hormones like steroid, vitamin D, retinoid and thyroid act through nuclear receptors. Insulin signals many body tissues whether the blood glucose is higher than necessary, so that cells take up excess glucose from the blood and convert it to glycogen and triacylglycerol. Low blood glucose is signaled by glucagon and tissues respond by producing glucose through glycogen breakdown and by oxidizing fats to reduce the use of glucose.