Unit 3

monomer/polymer chemistry 1

many biomolecules are large and complex, with thousands or millions of atoms 

many important biomolecules are made up of smaller building block molecules 

large biomolecules made using building block molecules called polumers 

the building block biomolecules are called monomers 

proteins - polymers made of amino acids 

    amino acids are monomers 

nucleic acids - polymers of nucleotides 

    nucleotides are monomers 

polysaccharides - polymers of sugar

    sugar molecule (often glucose) are monomers 

monomer structure 

monomers are made of a carbon skeleton with functional group(s) attatched 

functional froups are the sites wehre monomers attach to one another 

functional groups have only a few atoms 

monomers are bonded together at functional groups by a reaction called dehydration synthesis 

linked monomers can be broken apart between functional groups by reaction called hydrolysis 

monomers are building blocks 

polysaccharides, nucleic acids, and proteins are large biomolecules (polymers) made of smaller building block molecules (monomers) 

polysaccharides means "many sugars" and starch, a polysaccharide, is made up of long chains of thousands of glucose molecules 

DNA and RNA are the nucleic acids, and they are made of long chains of building block molecules called nucleotides 

proteins are also large biological molecules; proteins are polymers that are long chains of building block molecules; the building block molecules of proteins are the amino acids

like all monomers, each amino acid has its own important functional groups; in fact, they are called amino acids because their key functional groups are an amino group and a carboxylic acid group 

genetic information in action 

lactase is an enzyme (proteins) 

the lactase gene is a DNA sequence with the recipe for making the lactase enzyme 

lactase mRNA molecules are copies of the lactase gene, help cell make lactase

lactase molecules digest lactose molecules so they can be metabolized 

enzymes are proteins that do a job

lactase: shaping up right 

lactase molecules are proteins 

lactase molecules have the right shape to fit together with lactose molecules 

when a lactase molecule binds a lactose molecule, the lactase will digest the lactose molecule into its building blocks, the simple sugars glucose and galactose 

cells can use the simple sugars for energy 

protein conformation (shape)

proteins are molecules that gets things done

a protein molecule's ability to do its job depends on the protein's overall shape

for a protein to work, it has to attach to, or interact with other things; these interactions depend on the proper shape of the protein 

conformation is overall shape of proteins; i.e. tertiary or quaternary structure

correct conformation is required for protein molecules to function properly 

proper conformation required for: 

   enzymes to bind substrate 

    antibodies to bind antigens 

    bacterial exotoxins to poison cells 

    virus coat proteins to form the virus shell 

a protein molecules ability to do its job depends on the protein's overall shape; the overall shape of a protein is its conformation 

for a protein to work, it binds to or interacts with other substances; these interactions are totally dependent on proper protein shape 

changes in protein conformation will destroy the ability of protein to function 

protein denaturation 

a denatured protein no longer has its characteristic three dimensional shape 

denatured proteins are non-functional; they cannot work because lost conformation

people sometimes denature proteins on purpose; denatureation can be caused by cooking, autoclaving and many disinfectants and detergents 

denatutring proteins kills germs 

proteins and amino acids 

all proteins are made of building block molecules called amino acids 

because proteins are made of many smaller units (amino acids), they are called polymers 

because amino acids are the building blocks fo proteins, they are called monomers 

the terms building blocks and monomers have the same meaning 

protein facts 

all proteins made using universal set of twenty different amino acids 

average protein: about 300 amino acids 

there are very few proteins with < 50 amino acids

an E. coli cell can make about 5,000 different kinds of protein molecules

a typical bacterial cell contains hundreds of millions of protein molecules 

amino acids 

amino acids are so named because:

     they all have an amino group (-NH2)

    and a carboxylic acid group (-COOH)

these amino & carboxylic acid groups are the functional groups where amino acids are joined (dehydration synthesis) and where they are cleaved apart (hydrolysis) 

some amino acids also have functional groups located in the variable region 

amino acid structure 

central carbon atom: (a=alpha) 

a-carbon is bonded to four things

    lone hydrogen atom (-H) 

    carboxylic acid functional group (-COOH) 

   amino functional group (-NH2) 

variable group (-R) 

variable group (R group) is different in the different amino acids; it gives the amino acid its identity; page 44, R groups in blue 

peptide bond

the chemical bond between amino acids

the dehydration synthesis reaction forms peptide bonds between the amino acids 

    the carbon atom in the carboxyl group of one amino acid bonds to the nitrogen atom in the amino group of another amino acid 

    its dehydration synthesis reaction: the amino group loses -H; carboxyl group loses -OH

   the (-H) and the (-OH) combine to form H2O 

hydrolysis reactions

water molecules ionize in aqueous solutions; the equation is written: H2o -> H+ + OH-

so, in a volume of water, there is always a small but significant number of H+ and OH- 

in a hydrolysis reaction, H+ will bond with an atom on one functional group and OH- will bond to an atom of the other functional group and break the bond between the monomers

biochemical pathways 

many important metabolic processes have several steps; the steps are sequential 

each step in the process is a single chemical reaction catalyzed by a specific enzyme

all the steps, from start to finish, are known as a biochemical pathway

glycolysis, the kreb's cycle and the calvin cycle are examples of biochemical pathways

each step is catalyzed by a different enzyme 

    product of enzyme A is substrate for enzyme B

    product of enzyme B is substrate for enzyme C

example: making amino acids from sugar molecules, ammonia, and other chemicals; making fatty acids from sugar molecules

the enzymes involved are often held in place in order by being placed in cell membranes 

chemical energy

there is an exchange of energy whenever chemical reactions occur called chemical energy 

    endergonic reaction: requires energy; products have more chemical energy than reactants 

    exergonic reactions: release energy; products have less chemical energy than reactants 

in general, synthesis reactions require energy (endergonic) and decomposition reactions release energy (exergonic) 

anabolism and catabolism 

synthesis reactions- when atoms, ions, or molecules combine to make larger, more complex molecules; often referred to as an anabolic reaction in biology (anabolism) 

decomposition reactions- when larger, more complex molecules are broken into smaller atoms, ions, or molecules; often called catabolic reactions (catabolism)

catabolism

catabolic reactions release energy

catabolism usually refers to the breakdown of nutrient molecules like simple sugars, amino acids and fatty acids to generate energy 

digestion of a biological molecule like starch into its glucose building blocks isnt usually referred to as catabolism, even though it is a breakdown reaction; no energy generated 

metabolism

metabolism is the sum of all anabolic and catabolic reactions that occur in an organism

anabolic reactions require energy and are used to building larger molecules

catabolic reactions release energy by breaking down smaller energy molecules

catabolism plus anabolism is metabolism 

energy from catabolism used to make ATP

energy for anabolism comes from ATP

ATP and energy coupling

adenosine triphospate (ATP) is the main player in the transfer of energy in cells 

the chemical energy released by burning nutrient molecules like glucose is used to generate ATP molecules 

the cell uses ATP molecules to do work 

the energy released burning nutrients has to be trapped in ATP before it can be used 

ATP is an energy transfer intermediate 

ATP transfers energy from where it is released by burning nutrients, to where it is needed and used to do work within the cell 

    the energy released from nutrients is temporarily trapped in ATP molecules

    ATP molecules can deliver and release the trapped energy wehre it is needed to do work

this is called energy coupling 

ATP and cellular work

ATP contains energy in a form the cell can use for work. What work does a cell do?

    chemical work: sythesis of biomolecules like making proteins or copying nucleic acids

    transport work: moving things across plasma membrane, like food in and wastes out 

    mechanical work: moving chromosomes, spinning flagella, contracting muscles 

ATP provides useable energy for cell 

the ATP energy molecule is like a rechargeable battery

an ATP molecule has a packet of energy 

    it's charged 

this energy is released when bond between the 2nd and 3rd phosphates is hydrolyzed (broken)

when the 3rd phosphate is removed, then we don't have ATP anymore, but ADP (+P)

ADP no longer has the packet of energy

    it's discharged 

cell uses energy from breakdown of nutrients to recharge ADP (make it into ATP again) 

ATP isn't exactly like a battery

ATP goes directly from fully charged to fully discharged in a single reaction; nanoseconds 

ATP isn't used to store energy; ATP is made and used almost immediately; there is less than one minute between ATP formation and use 

ADP is recharged in a single reaction; this takes virtually no time at all 

ATP and energy

ATP provides energy that the cell can use 

ATP -> ADP + P is an exergonic reaction 

    the energy released can do work for the cell 

    ADP  has less energy than ATP 

ADP + P -> ATP is an endergonic reaction 

    the energy needed to power this comes from oxidizing (burning) nutrients like glucose 

    ATP has more energy than ADP 

ATP and metabolism 

energy is released in catabolic reactions 

energy is trapped briefly in ATP molecules 

ATP provides energy for anabolic reactions that make large molecules and for things like muscle contractions, flagella movement etc

energy released from catabolic reactions is used to make ATP, then ATP is used to drive anabolic reactions and other forms of work 

carbohydrate catabolism 

carbohydrate catabolism is a key source of energy for ATP production in living things

three types of carbohydrate catabolism 

    aerobic respiration: 

        glucose + O2 -> CO2 + H2O                    (38 ATP) 

    anearobic respiration: 

        glucose + not O2 -> CO2 + not H2O       (<38 ATP)

    fermentation: 

        glucose -> organic by-product                 (2 ATP) 

other energy nutrients 

amino acids can be broken down to provide energy (make ATP) for the cell 

fatty acids can be catabolized to provide energy to make ATP (from ADP and P) 

our focus: the catabolism of glucose 

polysaccharides like starch and glycogen are made of glucose; digested to form free glucose and glucose catabolized 

NADH: a high energy molecule

NADH is a key player in ATP production 

NADH: nicotinamide adenine dinucleotide, high energy (known as the reduced form) 

like ATP, NADH has a low energy form: NAD+ (known as the oxidized form) 

during some catabolic reactions, the energy released is used by the cell to make NADH from NAD+ and H (hydrogen) 

NADH has high energy electrons that will be used later to help make ATP 

aerobic cellular respiration 

often called aerobic respiration 

aka oxidative catabolism 

requires molecular oxygen (O2) 

purpose: generate useable energy (ATP) 

virtually all eukaryotes, some prokaryotes 

occurs in 3 stages 

    glycolysis / kreb's cycle / electron transport 

    each stage is a biochemical pathway 

gylcolysis 

first stage of aerobic respiration 

glycolysis is conversion of glucose (6-C) to two pyruvic acid (3-C) molecules 

biochemical pathway; 10 enzymatic steps 

glucose starting material; initial reactant 

pyruvic acid (pyruvate) is final product 

kreb's cycle

second stage of aerobic respiration 

pyruvate initial reactant; CO2 and H products 

biochemical pathway; 8 enzymatic steps 

remember 2 pyruvates per glucose enter 

    pyruvic acid converted to acetyl-CoA and enters Kreb’s cycle called prep phase 

    at the end of Kreb’s cycle all three carbons of pyruvate have been converted to CO2 

    cell gets one ATP and 5 NADH per pyruvate 

electron transport chain 

third stage in aerobic respiration 

biochemical pathway with proteins called electron carriers, as well as enzymes 

the ETC is where molecular oxygen is used; oxygen is the final electron acceptor 

the energy that was temporarily held in NADH is finally used 

cell gets 34 ATPs per glucose 

NADH from glycolysis and Kreb's cycle donates their high energy electrons to ETC 

electrons pass down the chain of electron carriers releasing energy 

released energy used to make ATP from ADP and P via ATP synthase (an enzyme) 

in last steps, electrons from ETC hook up with H+ and O2, forming H2O 

chemiosmosis: the parts 

the chemiosmotic theory explains how the electron transport chain generates ATP 

the electron carriers (7 total) are embedded in the membrane in sequential order 

some of the electron carriers are proton pumps that transport H+  across membrane

the symbol H+ represents the hydrogen ion 

ATP synthase is an enzyme that joins ADP and phosphate to make ATP molecules 

chemiosmosis: the process 

NADH gives its electrons to first electron carrier 

as electrons pass down the chain, they pass through carriers called proton pumps 

the proton pumps actively transport H+ to the other side of the membrane; the energy to do this comes from the flow of electrons down the ETC 

the H+ concentration on the other side of the membrane builds up to high levels; the uneven distribution of H+ is potential energy and is called the proton gradient (an H+ is a proton) 

the protons can flow back across the membrane only through special channels that contain the enzyme ATP synthase 

when H+ flows back across the membrane through the channels, ATP synthase uses the energy from the flow of protons to make ATP from ADP and phosphate 

anaerobic cellular respiration 

stages the same as in aerobic respiration 

    gylcolysis, Kreb's cycle, electron transport chain

ATP yield is variable, depends on bacteria and growth conditions; often approaches 38 ATP 

key difference is that an inorganic molecule other than O2 is final electron acceptor 

final electron acceptors: nitrate (NO3), sulfate (SO4), carbonate (CO3) and others 

some anaerobic bacteria 

different anaerobic bacteria use different chemicalsas final electron acceptors 

    Desulfovibrio: sulfate (SO4) is final electron acceptor, produce hydrogen sulfide (H2S) 

    Methanobacterium: carbonate (CO3) is final electron acceptor, produce methane (CH4) 

anaerobic bacteria do not need O2 to live and grow because they can generate ATP without using O2 (molecular oxygen) 

fermentation 

fermentation occurs in two stages 

    glycolysis: the conversion of glucose to pyruvate 

    fermentation: conversion of pyruvate to the fermentation end product 

many types of fermentation, named after the fermentation end product 

    lactic acid fermentation: lactic acid is fermentation end product 

    acetic acid fermentation: acetic acid is fermentation end product 

    alcoholic fermentation: ethanol is fermentation end product 

does not require oxygen 

can occur in the presence of oxygen 

yeilds two ATPs from each glucose 

not all bacteria are capable of fermentation 

regenerating NAD+ is purpose of converting pyruvate to fermentation end product 

NAD+ is converted to NADH during glycolysis and the cell has only so much NAD+ 

fermentation example 

brewers' yeast ferments glucose to ethanol 

stage 1 is glycolysis 

    glucose -> 2 pyruvate and 2 NAD+ +2 H -> 2 NADH

stage 2 is fermentation 

    2 pyruvate -> 2 ethanol + 2 CO2 and 2 NADH -> 2 NAD+ + 2 H 

purpose of stage 2 is to regenerate NAD+ 

purpose of stage 1 is to make the two ATPs 

catabolic energy production and O2

some key facts about molecular oxygen 

    some bacteria do not need O2: ones that are capable of anaerobic respiration; ones that are fermentative 

    oxygen is toxic to some bacteria: they lack enzymes to neutralize toxic forms; have only anaerobic pathways 

   some bacteria require oxygen: ones that have only the pathways of aerobic respiration; obligate aerobes 

   some bacteria, the facultative anaerobes and the aerotolerant anaerobes, can live with or without oxygen

    the air is 20% oxygen 

five categories based on O2 

obligate aerobes 

facultative anaerobes 

obligate anaerobes 

aerotolerant anerobes 

microaerophiles 

obligate aerobes 

require oxygen to live 

can only perform aerobic respiration 

Moraxella species (spp.) are aerobic; they cause conjuctivitis; the conjunctiva is the membrane covering the eyes and eyelids 

Neisseria spp. are aerobic bacteria; one species causes gonorrhea; a different species causes meningitis; meninges are membranes around brain and spinal cord 

facultative anaerobes 

all can perform aerobic respiration 

can switch to fermentation if oxygen is absent; so they can live without molecular oxygen 

growth is usually rapid in presence of oxygen and limited in its absence

Escherichia coli and many other enteric bacteria, like Klebsiella pneumoniae 

Saccharomyces cerevisiae bewer's yeast

a few facultative anaerobes switch from aerobic respiration to anaerobic respiration 

sometimes growth rate is not much affected by the absence of molecular oxygen 

usually the term facultative anaerobe refers to bacteria that can switch from aerobic respiration to fermentation 

obligate anaerobes 

oxygen is toxic to them; air kills them 

performs anaerobic respiration onlu 

some can form endospores to escape oxygen

Closstridium spp. can cause tetanus, botulism, food poisoning, gangrene; form endospores 

Bacteroides spp. non-endospore forming obligate anaerobes that inhabit digestive tract of humans and other animals 

aerotolerant anaerobes 

do not use molecular oxygen 

these microbes perform only fermentation 

grown equally well in presence or absence of molecular oxygen; but they grow slowly 

Streptococcus spp. are important human pathogens; strep throat and typical pneumonia 

Lactobacillus spp. used in dairy products, ferment carbohydrates to lactic acid

microaerophiles 

perform only aerobic respiration; use oxygen 

normal oxygen levels (20%) are harmful to these cells; usually grow best between 2%-5% oxygen 

often live in locations, or niches, within the host where oxygen levels are low 

Campylobacter is a leading cause of acute gastroenteritis; often contaminates poultry 

some reasons for growing bacteria 

vaccine production 

    grow pathogenic bacteria in culture, then kill them and use killed bacteria as a vaccine; old pertussis vaccine 

    grow bacteria in culture, then purify a part of the cell, like capsule, and use as a vaccine, pneumococcal pneumonia 

diagnosis of infection 

    traditional methods of identifying bacteria first require isolating and culturing bacteria in pure culture 

antibiotic production 

    nearly half of antibiotics are extracted from cultures of antibiotic-producing bacteria; tetracycline, erythromycin 

genetic engineering (biotechnology) 

    used to manufacture useful proteins 

   bacterial cells are given the gene (DNA) for useful protein and bacteria make the protein; human insulin

    useful protein purified from bacterial culture 

use in food and industry 

    to make many types of food, especially dairy foods

medical research

    to learn more about microbes and how they cause disease; how they live; how they can be controlled 

pure bacterial cultures

culture containing only one bacterial species 

pure cultures are essential for work involving bacteria, including traditional identification 

clinical samples may contain dozens of different bacteria; one must isolate pathogen and grow it in a pure culture for successful ID 

streak plate technique often used here 

culturing bacteria successfully 

proper physical conditions for growth 

    temperature 

    pH

    osmotic pressure 

        isotonic, hypertonic, hypotonic

proper chemical requirements 

    macronutrients 

   growth factors 

    trace elements 

laboratory conditions optimized for maximum growth 

temperature - for most clinical bacteria, it's near body temperature (98F or 37C) 

osmotic pressure- near isotonic; the solute concentrations inside and outside cell are nearly equal; growth media is near isotonic 

pH- the pH of culture media is near neutral; media generally has pH between 6 & 8; pH 7 is neutral 

bacterial growth 

bacterial growth really means multiplication 

bacteria reproduce by binary fission 

one cell becomes two, two becomes four, etc 

bacteria can reproduce in a short time; time between cycles is called generation time 

generation time for most bacteria is between twenty and sixty minutes 

m x 2^n = p; m = # of starting cells, n = # of generations, p = # of cells in population 

chemical growth requirements 

CHONPS - carbon, hydrogen, oxygen, nitrogen, phosphorus, and sulfur; called macronutrients; about 98% of the cell 

organic growth factors - pre-made amino acids, nucleotides, and vitamins; required for growth of some microbes 

trace elements - minerals and metals; calcium, zinc, magnesium 

macronutrients 

carbon: carbon atoms in carbon skeleton 

hydrogen: fills in needed bonds 

oxygen: two kinds are important 

    o2 from air: reactant in cellular respiration 

    o as in h2o; c6h12o6 

nitrogen: amino acids, nucleotides, and ATP 

phosphorus: nucleotides and phospholipids 

sulfur: in some amino acids (proteins) 

biomolecule breakdown 

proteins: contain COHNS 

starch and most carbohydrates: contain CHO 

nucleic acids: contain CHONP 

phospholipids: contain CHONP 

lipids: contain CHO

peptidogylcan: contains CHON 

culture media for bacteria 

complex media - aka general purpose media; made from natural products like plants or animals 

anaerobic media - for growing anaerobes; has chemicals that trap molecular oxygen 

selective media - selects for different types of bacteria; grows one type better than others 

differential media - differentiates between bacteria; the media (or the growth) will appear different if the bacteria growing in the media have different biochemical characteristics 

complex media 

made from natural products like animal, plant, or yeast extracts; peptones often added 

lots of CHONPS in the media, these elements are part of different biomolecules, like proteins, amino acids, and carbohydrates found in the extracts 

examples: nutrient broth, nutrient agar, tryptic soy broth, tryptic soy agar, brain heart infusion

complex media are for routine culturing of bacteria; aka general purpose media 

anaerobic media 

in addition to nutrients like shown above, anaerobic media contains chemicals that remove or soak up molecular oxygen (o2) 

the chemicals that remove molecular oxygen are called reducing agents 

it's oxygen free due to the reducing agent 

used to culture obligate anaerobes, which are killed by molecular oxygen; Clostridium 

selective media 

used to grow desired bacteria by inhibiting the growth of undesired bacteria 

used to separate (or select) desired type of bacteria from mixtures of bacteria 

most selective media will grow only gram positive or only gram negative bacteria 

selective media select for a particular kind of bacteria; media x selects for gram+, which means that g+ bacteria grow and g- bacteria dont 

examples of selective media

brilliant green agar- contains a dye that prevents peptidoglycan layering; this slows or prevents the growth of gram - positives; BGA selects for gram negative bacteria 

phenyl ethyl alcohol agar - contains PEA, which dissolves the outer membrane of the gram negative cell wall; prevents the growth of gram negatives; gram positives do grow; we say it selects for gram positive bacteria 

differential media 

will grow most, if not all, types of bacteria 

used to differentiate between species of bacteria based on bacterial biochemistry 

usually contain chemical indicators that change color in the presence of specific substances made during bacterial growth 

widely used in bacterial identification; used to determine specific biochemical characteristics of unknown bacteria 

example of differential media 

phenol red fermentation tubes 

    media contains a known carbohydrate, broth with non-carbohydrate nutrients, phenol red 

    phenol red is a pH indicator 

        red at pH 7 and turns yellow at acidic pH (below pH 5) 

    if bacterium growing in this broth ferments the carbohydrate to acid, the media will turn yellow 

    if bacterium can't ferment carbohydrate, it grows on the non-carbohydrate nutrients (peptones) present in the broth and the media stays red 

blood agar 

    this is a TSA based media that contains sheep red blood cells; media is reddish brown in color 

    some bacteria can lyse blood cells and degrade the hemoglobin (hg); hg makes blood red 

    these hemolytic bacteria will be surrounded by a tan colored halo on blood agar

    non hemolytic bacteria will not have the halo 

    blood agar differentiates between the two types 

combined selective and differential 

some media are selective and differential 

mannitol salt agar (MSA)

    salt (NaCl) in the media selects for salt tolerant bacteria; meaning that only bacteria that can tolerate high salt levels will grow 

    the media also has mannitol (a carbohydrate) and phenol red; if the bacteria ferments the mannitol, the pH drops and media will turn from pink to yellow; no fermentation, stays pink