Sri Raghavendra Educational Institutions Society (R)

(Approved by AICTE, Accredited by NAAC, Affiliated to VTU, Karnataka)

Sri Krishna Institute of Technology

www.skit.org.in

Title: BIOMOLECULES & THEIR APPLICATIONS

CO addressed:CO1- Understand the basic biological concepts through relevant

industrial applications and case studies.

Course: BIOLOGY FOR ENGINEERS(BBOK407)

Presented by: Mr. SATISH S

Department: Electronics & Communication Engineering

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Sri Krishna Institute of Technology

BIOMOLECULES AND ITS APPLICATIONS

SYLLABUS

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Carbohydrates (cellulose-based water filters, PHA and PLA as bioplastics)

Nucleic acids (DNA Vaccine for Rabies and RNA vaccines for Covid19, Forensics – DNA fingerprinting)

Proteins (Proteins as food – whey protein and meat analogs, Plant based proteins),

Lipids (biodiesel, cleaning agents/detergents)

Enzymes (glucose-oxidase in biosensors, lignolytic enzyme in bio- bleaching).

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BIOMOLECULES AND ITS APPLICATIONS

What is a biomolecule?

Biomolecule, also called biological molecule, any of numerous substances that are produced by cells and living organisms.

Biomolecules have a wide range of sizes and structures and perform a vast array of functions. The four major types of biomolecules are carbohydrates, lipids, nucleic acids, and proteins. Among biomolecules, nucleic acids, namely DNA and RNA, have the unique function of storing an organism’s genetic code—the sequence of nucleotides that determines the amino acid sequence of proteins, which are of critical importance to life on Earth.

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Carbohydrates, which are made up primarily of molecules containing atoms of carbon, hydrogen, and oxygen, are essential energy sources and structural components of all life, and they are among the most abundant biomolecules on Earth. They are built from four types of sugar units— monosaccharides, disaccharides, oligosaccharides, and polysaccharides.

Lipids, another key biomolecule of living organisms, fulfill a variety of roles, including serving as a source of stored energy and acting as chemical messengers. They also form membranes, which separate cells from their environments and compartmentalize the cell interior, giving rise to organelles, such as the nucleus and the mitochondrion, in higher (more complex) organisms. Examples include cytidine, uridine, adenosine, guanosine, and thymidine. Nucleosides that are phosphorylated become nucleotides. Apart from serving as a structural unit of nucleic acids, nucleotides may also serve as sources of chemical energy (e.g. adenosine triphosphate or ATP).

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MAIN APPLICATIONS OF

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CARBOHYDRATES:

What are carbohydrates?

Carbohydrates, or carbs, are sugar molecules. Along with proteins and fats, carbohydrates are one of three main nutrients found in foods and drinks. Body breaks down carbohydrates into glucose. Glucose, or blood sugar, is the main source of energy for your body's cells, tissues, and organs. Glucose can be used immediately or stored in the liver and muscles for later use.

There are three main types of carbohydrates:

1. SUGARS : Further classified into i) Natural ii) Starches iii) Fibers
2. Cellulose: a complex carbohydrate, or polysaccharide, consisting of 3,000 or more glucose units. It is extremely abundant, easily renewable, and biodegradable

Taking benefit of these advantages of cellulose, we have a best application of cellulose, that is,

Cellulose-based water filters.

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• Creating biobased membranes for micro- and ultrafiltration from cellulose nanofibrils (CNFs).
• Filters based on cellulose pulp fibers do usually have large pores that facilitate water percolation but they do not sufficiently remove bacteria through size exclusion
• Several groups have addressed this issue by incorporating antibacterial metal nanoparticles into cellulose-based water filters; both silver nanoparticles (AgNPs) and copper nanoparticles (CuNPs) are known to have good antibacterial effects.

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Both Gram-positive and Gram-negative bacteria have a negative net surface charge on the cell envelope, due to peptidoglycans, liposaccharides and proteins in the cell wall, and this makes their removal non- selective and efficient for most types of bacteria.

Methods used for the same are:

• LBL [Layer By Layer] MODIFICATION
• NITROGEN ANALYSIS
• SEM [ Scanning Electron Microscope]
• FLOW RATE FOR FREE FLOW FILTRATION
• BACTERIAL REMOVAL EFFICIENCY OF FILTRATION
• FILTRATION OF NATURAL WATER SAMPLES
• FLOURESCENCE MICROSCOPY

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Cellulose filter papers are versatile and diverse tools for microfiltration, that work by trapping particulates within a random matrix of cellulose fibers. Cellulose filter papers can be categorized as quantitative or qualitative, depending on their application.

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Cellulose based nanomaterials for water treatment

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APPLICATION 2:

Bioplastics are one type of plastic which can be generated from natural resources such as starches and vegetable oils. Bioplastics are basically classified as bio based and/or biodegradable

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The bioplastics are considered compostable if within 180 days, a complete microbial assimilation of the fragmented food source takes place in a compost environment.

Based upon this, we have PHA and PLA.

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PLA is both: biobased and biodegradable under industrial composting conditions (at a high temperature, around 58 °C). Because of its good mechanical properties, processability, renewability, and non-toxicity

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PHAs are a significant polymer family that are 100% bio-based and bio-degradable. PHAs are microbiologically produced polyesters that have tunable physical and mechanical properties

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STEPS IN PRODUCING BIOPLASTICS FROM PHA AND PLA

Nucleic Acids

Nucleic acids are biopolymers that play a crucial role in the storage and transfer of genetic information in all living organisms.

There are two types of nucleic acids:

• Deoxyribonucleic acid (DNA): DNA is the genetic material that carries the instructions for the development, functioning, and reproduction of all living organisms. DNA is a double- stranded helix structure composed of nucleotides, which consist of a sugar (deoxyribose), a phosphate group, and a nitrogenous base (adenine, guanine, cytosine, or thymine).

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• Ribonucleic acid (RNA): RNA is involved in the expression of the genetic information stored in DNA by carrying the message from the DNA to the ribosome, where it is used to build proteins. RNA is a single-stranded molecule composed of nucleotides, which consist of a sugar (ribose), a phosphate group, and a nitrogenous base (adenine, guanine, cytosine, or uracil).

Both DNA and RNA play essential roles in the functioning of cells and organisms, and their structures and interactions with other molecules are the basis for many biological processes such as replication, transcription, and translation.

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DNA VACCINE FOR RABIES:

• A DNA vaccine, using a pCl-neo plasmid encoding the glycoprotein gene of a Mexican isolate of rabies virus, was developed to induce long-lasting protective immunity against rabies virus in dogs.
• The worldwide incidence of rabies and high rates of therapy failure, despite availability of effective vaccines indicate the need for timely and improved prophylactic approaches.
• DNA vaccination based on optimized formulation of lysosome-targeted glycoprotein of the rabies virus provides potential platform for preventing and controlling rabies.

RNA VACCINES FOR COVID-19:

• The mRNA molecule is essentially a recipe, telling the cells of the body how to make the spike protein. COVID-19 mRNA vaccines are given by injection, usually into the muscle of the upper arm.
• After the protein piece is made, the cell breaks down the instructions and gets rid of them. The mRNA never enters the central part (nucleus) of the cell, which is where our DNA (genetic material) is found. Your DNA can't be altered by mRNA vaccines.
• The cell then displays the protein piece on its surface. Our immune system recognizes that the protein doesn't belong there and begins building an immune response and making antibodies

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DNA fingerprinting: also called DNA typing, DNA profiling, genetic fingerprinting, genotyping, or identity testing, in genetics, method of isolating and identifying variable elements within the base-pair sequence of DNA (deoxyribonucleic acid).

1. The procedure for creating a DNA fingerprint consists of first obtaining a sample of cells, such as skin, hair, or blood cells, which contain DNA.
2. The DNA is extracted from the cells and purified, the DNA was then cut at specific points along the strand with proteins known as restriction enzymes.
3. The enzymes produced fragments of varying lengths that were sorted by placing them on a gel and then subjecting the gel to an electric current (electrophoresis): the shorter the fragment, the more quickly it moved toward the positive pole (anode).
4. The sorted double-stranded DNA fragments were then subjected to a blotting technique in which they were split into single strands and transferred to a nylon sheet

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FORENSIC-DNA FINGERPRINTING

5. The fragments underwent autoradiography in which they were exposed to DNA probes

6. The resultant pattern of marks could then be analyzed

7. The DNA testing process is comprised of four main steps, including

extraction, quantitation, amplification, and capillary electrophoresis

PROTEINS 

Protein is found throughout the body—in muscle, bone, skin, hair, and virtually every other body part or tissue. It makes up the enzymes that power many chemical reactions and the hemoglobin that carries oxygen in your blood.

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• Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues
• Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, responding to stimuli, providing structure to cells and organisms, and transporting molecules from one location to another.

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PROTEIN AS FOOD:

• Protein is a key part of any diet. The average person needs about 7 grams of protein every day for every 20 pounds of body weight
• Animal-based foods (meat, poultry, fish, eggs, and dairy foods) tend to be good sources of complete protein, while plant-based foods (fruits, vegetables, grains, nuts, and seeds) often lack one or more essential amino acid.

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Whey protein

It is a mixture of proteins isolated from whey, the liquid material created as a by- product of cheese production

• The proteins consist of α-lactalbumin, β-lactoglobulin, serum albumin and immunoglobulins. Glycomacropeptide also makes up the third largest component but is not a protein
• Whey protein is commonly marketed as a protein supplement, and various health claims have been attributed to it
• Whey is left over when milk is coagulated during the process of cheese production, and contains everything that is soluble from milk after the pH is dropped to 4.6 during the coagulation process.
• It is a 5% solution of lactose in water and contains the water soluble proteins of milk as well as some lipid content
• The primary usage of whey protein supplements is for muscle growth and development. Eating whey protein supplements before exercise will not assist athletic performance, but it will enhance the body's protein recovery and synthesis after exercise because it increases the free amino acids in the body's free amino acid pool.

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Meat analogues

• A meat alternative or meat substitute(Also called plant-based meat, mock meat or fake meat) is a food product made from vegetarian or vegan ingredients, eaten as a replacement for meat.

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• High moisture extrusion cooking enables the production of fresh, premium meat analogues that are texturally like muscle meat from plant or animal proteins. The appearance and eating sensation are similar to cooked meat while high protein content offers a similar nutritional value.

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• Meat analogues, can be defined as products that mimic meat in its functionality, bearing similar appearance, texture, and sensory attributes to meat. Production of meat analogues has been on the increase, targeted at satisfying consumers’ desire for indulgent, healthy, low environmental impact, and ethical meat substitutes

Plant Based Proteins

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Currently, available marketed meat analog products are plant-based meat in which the quality (i.e., texture and taste) are similar to the conventional meat.

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The ingredients used are mainly soy proteins with novel ingredients added, such as mycoprotein and soy leghemoglobin.

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Plant protein is simply a meaningful food source of protein which is from plants. This group can include pulses, tofu, soya, tempeh, seitan, nuts, seeds, certain grains and even peas.

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Pulses are a large group of plants, which include chickpeas, lentils, beans (such as black, kidney and adzuki beans) and split peas.

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Plant proteins are highly nutritious – not only as good sources of protein, but also because they provide other nutrients such as fibre, vitamins and minerals.

Our intake of fibre tends to be too low, however by incorporating certain plant proteins into your diet, such as pulses, peas and nuts, you can easily boost your fibre intake.

Consumer demand for plant protein-based products is high and expected to grow considerably in the next decade

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Factors contributing to the rise in popularity of plant proteins include:

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1)potential health benefits associated with increased intake of plant-based diets

2) consumer concerns regarding adverse health effects of consuming diets high in animal protein (e.g., increased saturated fat)

3) increased consumer recognition of the need to improve the environmental sustainability of food production

4) ethical issues regarding the treatment of animals

5) General consumer view of protein as a “positive” nutrient (more is better). While there are healthand physical function benefits of diets higher in plant-based protein, the nutritional quality of plant proteins may be inferior in some respects relative to animal proteins.

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ENZYMES

Our bodies naturally produce enzymes.

The six kinds of enzymes are hydrolases, oxidoreductases, lyases, transferases, ligases and isomerases.

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Enzymes perform the critical task of lowering a reaction's activation energy—that is, the amount of energy that must be put in for the reaction to begin.

Enzymes work by binding to reactant molecules and holding them in such a way that the chemical bond-breaking and bond-forming processes take place more readily

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Additionally, the high specificity of enzymes enhances the ability to detect lower analyte concentration limits. So enzymes are used in biosensors.

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Biosensors are employed in applications such as disease monitoring, drug discovery, and detection of pollutants, disease-causing micro-organisms and markers that are indicators of a disease in bodily fluids (blood, urine, saliva, sweat).

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Various types of biosensors being used are enzyme-based, tissue-based, immunosensors, DNA biosensors, and thermal and piezoelectric biosensors. There are wide variety of enzymes used in biosensors. One such enzyme is glucose oxidase, mainly in amperometric glucose biosensor.

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GLUCOSE OXIDASE IN BIOSENSORS:

Glucose oxidase (GOx) is widely used enzyme in glucose biosensor due to its better stability and relatively inexpensive. GOx catalyses the redox reaction and transfer electrons from enzyme active sites to electrode for glucose level analysis in blood samples.

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Quantification of glucose can be achieved based on either the detection of the hydrogen peroxide produced or the oxygen consumed

LIGNOLYTIC ENZYME IN BIO BLEACHING:

• Ligninolytic enzymes play a key role in degradation and detoxification of lignocellulosic waste in environment. The major ligninolytic enzymes are laccase, lignin peroxidase, manganese peroxidase, and versatile peroxidase.
• Ligninolytic fungi and enzymes (i.e., laccase, manganese peroxidase, and lignin peroxidase) have been applied recently in the production of second-generation biofuels.
• White-rot fungi are the main producers of lignin-oxidizing enzymes

• These fungi secrete a number of oxidative enzymes and some hitherto unknown substances (mediators) into their environment together effecting a slow but continuous degradation.
• The most important lignin-oxidizing enzymes are lignin peroxidases, manganese peroxidases and laccases.
• Lignin peroxidase and manganese peroxidase appear to constitute a major component of the ligninolytic system

LIPIDS

• Lipids, are a broad group of naturally occurring molecules which includes fats, waxes, sterols, fat-soluble vitamins (such as vitamins A, D, E and K), monoglycerides, diglycerides, phospholipids
• The functions of lipids include storing energy, signaling, and acting as structural components of cell membranes
• Lipids have applications in the cosmetic and food industries, and in nanotechnology.
• Lipids may be broadly defined as hydrophobic or amphiphilic small molecules
• The amphiphilic nature of some lipids allows them to form structures such as vesicles, multilamellar/unilamellar liposomes, or membranes in an aqueous environment.
• Biological lipids originate entirely from two distinct types of biochemical subunits or "building-blocks": ketoacyl and isoprene groups

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• Lipids may be divided into eight categories:

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• Fatty acyls, glycerolipids, glycerophospholipids, sphingolipids, saccharolipids, and polyketides (derived from condensation of ketoacyl subunits)

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• Sterol lipids and Prenol lipids (derived from condensation of isoprene subunits).

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• The formation of lipid bilayers is an energetically preferred process when the glycerophospholipids described above are in an aqueous environment.
• This is known as the hydrophobic effect.
• In an aqueous system, the polar heads of lipids align towards the polar, aqueous environment, while the hydrophobic tails minimize their contact with water and tend to cluster together, forming a vesicle;
• Depending on the concentration of the lipid, this biophysical interaction may result in the formation of Lipid bilayers

Classification of LIPIDS

Some of the applications are:

• Within the body, lipids function as an energy reserve, regulate hormones, transmit nerve impulses, cushion vital organs, and transport fat-soluble nutrients.

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• Fat in food serves as an energy source with high caloric density, adds texture and taste, and contributes to satiety.

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• Lipid obtained from food waste was used as a potential feedstock for biodiesel production using both a chemical catalyst and a biocatalyst. Base (KOH) catalyzed transesterification of the lipid allowed a 100% conversion of biodiesel in 2 h; whereas, Novozyme-435 yielded 90% biodiesel in 24 h. So lipids are having a main application in bidiesel production.

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• Biodiesel demonstrates an animal fat-based or vegetable oil diesel fuel, including long-chain alkyl (methyl, ethyl, or propyl) esters. Biodiesel is generally made by esterifying lipids (e.g., soybean oil, vegetable oil, and animal fat (tallow)) with an alcohol generating fatty acid esters. Biodiesel is suggested to be utilized in standard diesel engines and is thus well defined from the vegetable and waste oils used to fuel converted diesel engines. Biodiesel can be used singly or blended with gasoline in any proportions. Biodiesel blends can also be utilized as heating oil

Lipids also has an interesting application in cleaning agents.

• The hydrophobic end of the phospholipid bilayer stays away from the water. This avoids the dissolution of cell membrane in water. But the detergent can bind to the hydrophobic end of the cell membrane and form a solution with water, thus breaking the cell membrane barrier.
• Detergent monomers solubilize membrane proteins by partitioning into the membrane bilayer. With increasing amounts of detergents, membranes undergo various stages of solubilization. The initial stage is lysis or rupture of the membrane.

While lipids also have the same general structure as detergents—a polar hydrophilic head group and a nonpolar hydrophobic tail— lipids differ from detergents in the shape of the monomers, in the type of aggregates formed in solution, and in the concentration range required for aggregation