Reflecting back on the course, what are three major themes you would identify that connect the various topics discussed in this course – how are they connected to more than one topic, and how do they connect with what you knew before this course? What knowledge have you gained with regards to these three themes you have identified?

Various themes are present in Biochemistry that make it possible to see many connections with other biology and chemistry principles.  Biochemistry bridges these two disciplines in many ways; in explaining how principles of both disciplines govern the structure and function of organisms.  Reflecting back on the course and my studies, it has become apparent that the purpose of Biochemistry is to understand this bridge and how chemistry underlies and explains all biological processes (and how these individual chemical reactions might be targeted and altered to the advantage of the organism and to the disadvantage of disease).  Specific to this ”biochemical bridge”, three major themes have resounded through my  study of biochemistry and biochemical processes in the body:  chemical pathways and how they are interwoven, catalysis and control, and energy conversion.

Chemical Pathways & How they are Interwoven

An organism’s processes are comprised of chemical pathways that consist of many series of chemical reactions and steps that involve various molecules that lead to the creation of structures and functions vital to the organism.  These pathways are also intricately interwoven in a way that allows that organism to change quickly and efficiently respond to its internal and external environment.  Various pathways or steps in pathways can turn "on" or "off' other processes.  For example, DNA replication in eukaryotes is initiated by a complex of proteins that bind to replicators.  Some of these proteins; appropriately named replication licensing factors (RLFs); are cytosolic, meaning they are located outside of the nucleus, in the cytosol, and are separated from the DNA by the nuclear membrane.  During cell mitosis the nuclear membrane dissolves.  This makes it possible for the cytosolic RLFs to gain access to binding to the origin recognition complex and DNA and begin replication.  Thus, the pathway for replication in eukaryotes is intricately linked with and dependent on the pathways of cell mitosis.  Induction of one set of pathways makes the other possible.  We also see many examples of interwoven pathways when we study the many pathways of metabolism, as many of the intermediates created in the chemical reactions of the catabolic pathways, are also used in anabolic pathways to build the molecules of life.  In this sense, the direction and which pathways are activated are adjusted based on the immediate needs of the organism.  In conclusion, the ways in which these chemical pathways are linked together and affect one another are vital to the proper functioning of the organism.

Catalysis and Control

Another very important theme of this “biochemical bridge” is catalysis.  So many of the reactions in various pathways are nonspontaneous and even those ones that are spontaneous, if left uncatalyzed, would happen far too slowly to prove any use to an organism that needs to continually adjust to rapid changes and stimuli.  As we have seen in every reaction in glycolysis, the citric acid cycle, electron transport chain, β oxidation, etc., an enzyme or enzyme complex is involved in every step, catalyzing each chemical reaction and providing chemical control points that allow various pathways to be activated and inhibited, depending on the current needs of the organism.  Previous to studying biochemistry, I had encountered the topic of enzymes in relation to their role in digestion and how they function in catabolic processes.  Continuing with this theme, we have studied the enzymes involved in the breakdown of glucose to pyruvate.  On the flip side, we have also studied the enzymes that catalyze gluconeogenesis reactions- those anabolic reactions that respond to the energy needs of the organism being fulfilled, so that glucose can be resynthesized, stored and used later, when energy supplies are low.  We have also seen how these enzymes can be turned on or off (by other enzymes!) and, thus, function as control points for the pathways that they catalyze.   Enzymes are essential to life processes.

Energy Conversion

Lastly, the theme of energy conversion is ever present throughout all biochemistry principles.  This energy conversion is the main theme in our studies of the various metabolic cycles.  Energy is created from each of the cycles and used to drive anabolic metabolism.  Each and every biomolecule requires energy in its production, every process that we have previously studied in anatomy and physiology involves use of energy converted and then transferred within the body.  Even the very processes of energy conversion require energy already converted and created by the body.  To be specific, in the glycolytic cycle, 2 ATP molecules are required in the series of reactions that convert a glucose molecule to 2 pyruvate molecules, a process which produces 4 ATP molecules for an energy-yield of +2 ATP.  We need ATP to make ATP! and other biomolecules too. 

How would you explain the connection between glucose entering the body and energy created by the body to a friend, using your new biochemistry knowledge?

Glucose is a food source of energy for the body.  It is a simple sugar and once ingested, it enters the bloodstream through the digestive system and is distributed to the cells of the body.  These cells need to take in the glucose and access the energy of the molecule and use it to create the form of energy that is usable by the body- a molecule called ATP.  This molecule “traps” energy to be tapped in 3 high-energy bonds.  The “TP” stands for triphosphate:  Each phosphate bond contains a large amount of energy that can be accessed and utilized by the cell when broken and each ATP molecule contains three of these high energy bonds.

Here is a brief glimpse of the path that glucose takes in the creation of this energy housing molecule:  Once within a cell’s cytoplasm, a glucose molecule is broken down into two smaller molecules through a series of chemical reactions that yield 2 high energy ATP.  These reactions take place without molecular oxygen and, thus, are labeled as anaerobic.  This pathway, called glycolysis, is an important pathway for the body for two major reasons: First, it produces the form of molecule necessary to continue on with other energy producing pathways, and secondly, it can produce energy for the body when oxygen levels are low.  The molecules produced in glycolysis are now in a form that can enter into the mitochondria: the major site of energy production in the cell.  Within the mitochondria, the molecules enter the Citric Acid Cycle: a cyclic path of reactions that produces more ATP energy molecules as well as electron carriers.  Electron carriers are molecules that carry electrons transferred from intermediate molecules of the pathway to a final pathway called the electron transport chain.  This final pathway uses the shuffling of these electrons to the ultimate electron acceptor; oxygen; in a series of chemical reactions to pump protons across a membrane and harness the energy in many more high energy ATP bonds that can be utilized by the body. 

Important to note is that this journey that glucose takes through these energy-producing pathways is not a one-way street.  Many times the body calls for energy to be stored and this flow will stop and energy storage pathways will be utilized.  These are pathways that bind glucose molecules together and store them for times when there is a high-energy demand, like when you go for a run or engage in some other kind of exercise or hard work that calls for large amounts energy quickly.  Also important to remember is that ATP is used by the body to drive anabolic processes as well: the building up of molecules needed by the body.  Many of the chemical intermediates within the energy-producing pathways are also used in molecule building pathways.  In short, our body’s energy production, energy usage and energy storage pathways are complex and interwoven and quickly reactive to the immediate needs of the body as they change from one moment to the next.  Glucose gets pulled apart, put back together and transformed in many complex chemical reactions to meet the energy producing and energy storage demands of the body from one moment to the next.

What knowledge have you connected with past knowledge?

Ah, yes- your eyes are correct; your synapses are firing in the right directions; you are not mistaken:  You have read this question before- precisely two entries back!  Since this is a repeat question, I have chosen to approach my answer a bit differently this time:  I will begin my answer with a bit of an editorial angle before I label several specific connections that biochemistry has made for me with past studies.  However, before I do so, it is noteworthy to point out that this same question could be used for each week’s blog entry for this class, for biochemistry is full of connections to knowledge I have previously gained in past classes and explanations of "why and how".

On the Editorial Side: 

I simply love the Biochemical Connections boxes in our text:   As a student of medical and health sciences, these lovely little half-page snippets have become a lifeline for me, as they often connect complicated sequences of biochemical steps to the world I am working to enter into, through explanations of how these processes, or problems with the processes, cause various diseases or genetic differences.  Others explain the basic physiology of which I have previously studied in further detail and provide a thoughtful pause to make these connections of how chemistry basics are the fundamental basis behind what we are studying.  Taken together, each of these Biochemical Connections boxes is analogous to an island oasis in the midst of an ocean of protein abbreviations, nucleotide sequences and nucleophilic attacks!

Recent Connections:

In the most recent chapters of study, we have been immersed in the world of DNA.  Namely, the study of the biochemical processes of DNA replication, transcription and translation.  Thus, I felt it most appropriate to focus on a discussion of specific examples of connections I have made to past knowledge in these areas.

Connecting back to Microbiology:

In our recent study of DNA replication, we learned about the processes of proofreading and repair mechanisms in prokaryotic DNA.  In addition to errors made in replication to cause mutation, other mutagens can also have an effect on damaging DNA sequence and structure.  In this section I immediately connected back to my study of Microbiology when we learned about these various sources of mutagens, specifically UV light’s ability to cause thymine dimers; the formation of π electron bonds from the 5^th C and 6^th C of two adjacent thymine bases, which can alter the shape of the DNA molecule and cause errors in transcription and replication.  I understood this in my previous study, but only on a very high level.  Now, with my understanding of Organic Chemistry and Biochemistry, I can understand the nature of the bond that is formed and why this alteration in the three dimensional structure of the DNA molecule has such a drastic effect on the replication and transcription processes.  This connection also extends to the processes of repair mechanism and the need for recognition of the “correct strand”.  I distinctly remember learning in Microbiology about how prokaryotic DNA use methylation to flag the parent strand.  My understanding of this process stopped here at the time.  Now my understanding is much more complete, as I can connect back to Organic Chemistry and to Biochemistry and know that the adenine bases of the parent strands are methylated (meaning that -CH₃ is added) before replication, allowing enzymes to distinguish the parent strand (one without the mistake) from the newly synthesized stand (one with the mistake) and make the correction accordingly.

Connecting back to Anatomy & Physiology:

Another example I will give demonstrates connections between recently learned biochemistry concepts with those learned in Anatomy and Physiology.  This example involves the transcription factor cyclic-AMP-response-element-binding protein (CREB) which, in a neuron, activates genes associated with synapse-strengthening proteins that influence the development of long-term memories.  Reading about CREB and its role in the neuron provided a long overdue explanation for what those calcium ions were doing during depolarization when the calcium channels are opened:  They were activating enzyme pathways that ultimately activated CREB which activated the appropriate genes for the appropriate synapse-strengthening protiens!  I found this connection very interesting, as it provided a very specific example of the effect that transcription factors have on transcription and gene expression and connected with the broader level understanding of neurological function that I studied in Anatomy and Physiology.  In this sense (and, of course, many more examples that are like it) biochemistry has provided me with the explanation of "why and how" and the details that were missing in some of my prior studies.  This missing “why and how” have often times plagued me, as I have always thought it would be easier to understand the structure and function of the nervous system (and other biological systems) if I just had more detail to understand the underlying processes.

Find an interesting Biochemistry Website

http://themedicalbiochemistrypage.org/index.php

This website is a teaching website geared towards the medical and health sciences student that provides an approach to the biochemical view of the normal human physiological processes as well as those that are involved in disease.  For example, the reader can access the page on Glycolysis: Regulating Blood Glucose and learn about the normal biochemical process involved in the basic digestion of starches and sugars, the oxidation of glucose for energy and the process of blood glucose level regulation.  From that same home page, the reader may access information on diabetes mellitis; a disease of blood glucose regulation; the different possible types and an explanation of the pathophysiology involved.

The medical biochemistry website is a great tool for biochemistry connections for those entering the medical field and studying the health sciences.  It is easy to use and the topics covered are conveniently listed on the front page.  The majority of the site's information is accessible without a paid subscription.  Glossaries for medical/clinical terms and abbreviations are provided for the student and a clinical lab data section offers normal value ranges for blood analysis data.

What knowledge have you connected with past knowledge?

Biochemistry, by its very definition, bridges together various scientific disciplines.  Put simply: biochemistry bridges the study of the physical science of chemistry that explores the properties, interactions and energy between particles of matter with biology and its studies of living organisms and vital processes.  For a student with a fair amount of studying in both areas, biochemistry offers a chance to reflect and make many connections to concepts learned before, and to see how the scientific disciplines are bridged together, for a more complete understanding.

Thus far in this course, the most prominent high level connection I’ve made goes back to Anatomy and Physiology:  form always follows function.  Throughout our study of Human Anatomy and Physiology this concept is ground into our minds and remains the guiding principle for the study of all systems of the human body, from the high level of body systems, right down to the microscopic human cell.  We and our many parts are designed to carry out tasks in the most efficient manner to ensure the propagation and survival of the human species.  Biochemistry continues our studies with this same guiding principle, but at a closer angle- one that looks microscopically at the human cell and its functions and goes even farther to study the structure and interactions of the molecules that make up human tissue.  We study the structure of the various molecules involved, how they interact with each other and what function that has to human life.

For example, we have looked closely at hydrogen bonds between water molecules.  This is a simple way to view and understand these interactions that we then viewed in a much more complex protein structure.  We learned that these bonds are very important in determining the secondary structure of proteins; they are what hold the protein chain in helical arrangement, or pleated sheets, when the chain doubles back on itself or between two chains.  We have seen that the structure of these proteins is imperative to their function and the misfolding of the chains can cause these molecules to lose basic properties that allow their biological function:  for example they may misfold in a way that disrupts hydrophobic interactions, exposing the hydrophobic portions of the molecule which ultimately causes them to lose their solubility in the aqueous confines of the cell.  Aggregating together, they form a harmful substance, as is the case with the plaques associated with Alzheimer’s Disease.  Thus, hydrogen bonds, hydrophobic interactions and other inter-/intra-molecular interactions at play in the protein structure show a great example of how changing the structure and chemical interaction at the molecular level can have devastating impact on the function of anatomy at the systemic level that is studied in Human Anatomy and Physiology.

Another example of connection through biochemistry topics is that of enzymes.  In general chemistry we learned about catalysis and its effect on the free energy of activation.  In biochemistry, we are taking this chemical knowledge and seeing how it applies to proteins that we studied at a very high level in Anatomy and Physiology: enzymes.  We are now able to see the chemistry behind the enzymatic activity that is so incredibly vital to life processes and understand how and to what extent enzymes catalyze vital reactions.  The chemical structure and interactions at the enzyme active site give us a clear “how” to the process that we only touch upon in A&P.  The chemical structure or form shows us the “how” enzymes work, and the interactions and free energy of activation shows us the “why” these proteins provide the vital functions they do to carry out the processes and reactions of life that would otherwise occur way too slowly.

Through its connection of chemistry and organic chemistry fundamentals with their application to the molecules of life and function, biochemistry shows us how it all comes together, making sense of a multitude of previously introduced concepts.  I look forward to being able to see the rest of the connections that will unfold throughout this course.

Find a protein using PDB explorer- describe your protein, including what disease state or other real-world application it has.

Protein:  3DSF 

Anti-Osteopontin Antibody 23C3 in Complex with W43A Mutated Epitope Peptide

This protein is an antibody associated with an extracellular linking protein called osteopontin that has been linked to the auto-immune disease Rheumatoid Arthritis.  This antibody-eptitope peptide complex is the foundation of hopeful treatment for patients suffering with this auto-immune condition that consists of inflammation and damage of the body’s synovial joints by the immune system.

Patients with Rheumatoid Arthritis have greater than normal amounts of osteopontin present in their synovial tissue.  It has been shown experimentally that osteopontin plays a role in the pathogenesis of RA: one that recruits inflammatory cells through chemotactic action, bringing more immune cells to the synovium, increasing inflammation and ultimately causing more damage to the joints.  The use of this antibody/epitope peptide protein complex may offer therapeutic relief for RA patients in the development of drug treatments based on the binding site to osteopontin, to block its chemotactic activities and reduce T-cell responses, which in turn will decrease the amount of inflammatory cells and damage to the joints.  Inclusion of the specific epitope offers an additional range of possibilities for the development for new drugs to treat RA disease.

Primary Structure:

A total of 441 residues are found in this protein complex.

Secondary Structure:

This antibody/epitope peptide complex consists of mainly beta sheets with a small percentage of helices and a short peptide from the osteopontin protein.

Tertiary Structure:

The forces and interactions involved in 3DSF determine its structure as a globular protein.

Quarternary Structure:

3 chains make this protein complex a trimer.  Typically antibodies exist as tetramers, containing 4 very flexible chains: two long, heavy chains and two short, light chains.  However, the flexible nature of these globular proteins makes it extremely difficult to study them intact.  The specific structure for 3DSF contains a heavy chain (H), a light chain (L) and a peptide from osteopontin:

Chain H:  Fab fragment of anti-osteopontin antibody 23C3, Heavy chain

                                5% helical (4 helices; 12 residues)

                                48% beta sheet (21 strands; 104 residues)

Chain L:  Fab fragment of anti-osteopontin antibody 23C3, light chain

                                3% helical (2 helices; 7 residues)

                                49% beta sheet (21 strands; 106 residues)

Chain P:  Peptide from osteopontin (12 residues)

Much of the immune response and attack of the synovium in Rheumatoid Arthritis is still not completely understood and medications that have historically been used to treat this autoimmune condition can be harsh and often have many undesirable side effects.  Recently there has been a surge of research into the mechanisms involved in the pathogenesis of RA and other autoimmune diseases, offering a promise of more effective biological treatment.  It is clear that osteopontin plays a role in RA and several other autoimmune conditions.  The development of this protein complex may offer new potentially more effective and less harsh therapeutic possibilities to those who suffer with RA.

For additonal information:

http://www.pdb.org/pdb/explore/explore.do?structureId=3DSF

http://onlinelibrary.wiley.com/doi/10.1002/art.27603/pdf

http://www.pdb.org/pdb/101/motm.do?momID=21

What is Biochemistry, and how does it differ from the fields of Genetics, Biology, Chemistry and Molecular Biology?

Biochemistry is the study of the chemical structures and functions of the molecules of life, the vital processes occurring in living organisms and the roles of biomolecules in these processes.  Biochemistry provides a view of biomolecules at the chemical level and considers their chemical structure and behavior as determining their functions in life processes.  It is a scientific discipline largely concerned with metabolism: the catabolic processes employed to extract energy and the anabolic processes to build the vital molecules of life.  It is a discipline of biological and chemical sciences that intricately overlaps multiple scientific disciplines, including genetics, biology, chemistry and molecular biology.

Biochemistry is a branch of biology; the science that studies living organisms and vital processes; as it studies the very molecules of life.  However, it branches off from biology with its consideration of the chemistry of the molecules of life.  Biochemistry takes into account the chemical structure of biomolecules and considers the impact of this structure and chemical interactions in its study of their function and processes.  The physical science of chemistry focuses on the structure, properties and interactions of all particles of matter and the energy between them.  Thus, from this perspective, biochemistry is a discipline of chemistry also, differentiating from it by its roots in biology and its specific focus on biological matter and processes.

Genetics; the science of genes, heredity and variation in living organisms; is also very closely linked to biochemistry.  The structure and function of the biomolecules DNA and RNA are considered heavily in biochemistry, as they carry the code for construction of other biomolecules.  However, genetics takes a different path in its study, focusing mainly on the structure of the gene in the context of an organism, how that gene is expressed and ways in which its expression may be halted or altered.  This is a higher level view than the biochemistry platform, which studies these macromolecules and their structures for the purpose of understanding their function to the biological processes of an organism.  Biochemistry is more concerned with what proteins these molecules code for and create, the process by which they accomplish this, and what function these molecules serve in basic biological function.

The “lines” defining the different areas of biochemistry and molecular biology appear to be even more blurred as molecular biology also takes a close look at biological activity at the molecular level and studies the interactions of the systems of cellular function.  Like biochemistry, molecular biology considers the structure of the molecules of life and how they function in the biological processes that are necessary to life.  However, its focus gravitates more to the study of genetic origin, transcription of genetic material, and its translation to the molecules of life and their cellular functions, while biochemistry focuses more on nutrition, metabolism and biological functions at the molecular level.  Molecular biology has its roots firmly grounded in biochemistry and genetics and has rapidly become any area of great interest and study; one that is so vast that it has required a distinction from other disciplines as its own entity and area of study.

One must note that defining the differences between these disciplines may be essential to understanding their functions and purpose in the scientific world, however, each of these disciplines is interwoven with the next, each reaching for further understanding as it pulls knowledge and connection from the others.  In many ways, biochemistry, genetics and molecular biology have no concrete defined lines between them and overlap greatly.  Perhaps the best way to understand it is by the concept of perspective- genetics coming from the perspective of understanding the gene and its expression, biochemistry from the perspective of studying the structure of biomolecules and how this determines their function in biological processes and metabolic processes, and molecular biology from the perspective of studying the processes of transcription and translation of genetic material into proteins and the molecules of life.  Each perspective offers more discovery and, when taken together, a more complete understanding of the very structure and function of life.