Structural Biochemistry/Cell Organelles/Mitochondria

Function

Mitochondria seen through an electron microscope

The purpose of the mitochondria in the eukaryote is to provide cellular respiration to the cell. The endosymbiotic theory asserts that the mitochondria came to be part of the eukaryote over time through a symbiotic relationship. The mitochondria consists of two membranes, the inner membrane and the outer membrane. It is speculated that the outer membrane came about when its ancestor was engulfed by the host celled via endocytosis, giving it a membrane in addition to the one the mitochondria ancestor already had. This endosymbiont theory would also explain why the mitochondria had its own DNA and why this DNA is circular. For some amino acids the genetic code of the mitochondria differ slightly from that of the nucleus (and the rest of the cell). The mitochondria's energy from respiration is stored in an ion gradient across the organelle's double membrane, known as the "Mitchell Hypothesis".

Mitochondrial Biogenesis: It’s a metabolic adaptation where mitochondrial mass increases which allow for conduction of glycolysis, oxidative phosphorylation, and ultimately result in higher mitochondrial metabolic capacity. With greater capacity to synthesize and transport fuels to mitochondria, the metabolic response would be faster, which can be beneficial for athletes during exercise. However, this improvement requires exercise and training.

Inner Mitochondria & Matrix

Inside the deepest compartments of the mitochondria is the mitochondrian matrix. It is in the matrix that cellular respiration occurs, where pyruvate (a product of glycolysis in the cytosol) is converted to Carbon Dioxide and water. The matrix is the site of the the citric acid cycle, whereby the electron transport chain is used to setup a proton gradient between the inner and outer membrane of the mitochondria, known as the inter membrane space. The protons in the inter membrane space accumulate to a point that the concentration gradient causes the protons to flow back into the matrix.

It is the inner membrane that is studded with the proteins necessary for the electron transport chain, such as the cytochrome electron shuttles. Upon reentering the matrix, the H⁺ go through ATP synthase, which in turns powers the synthase to phosphorylate adenosine diphosphate (ADP) to adenosine triphosphate (ATP). The ATP can be used later on to be coupled with thermodynamically unfavorable reactions to allow those chemical reactions to proceed. The inner membrane is folded and convoluted which allows for a greater surface area to utulize for the electron transport chain. These convolutions are what make up the cristae.

Interestingly enough the matrix of the mitochondria are one of the few locations outside of the nucleus where genetic information can be found in the cell. Mitochondrial DNA is similar in appearance to that of bacterial DNA due to its circular shape. The matrix is also known to house tRNA and ribosomes, which further solidifies the theory that the mitochondria entered the ancestral eukaryotic cell as single celled organism.

Outer Membrane

The outer mitochondria membrane consist of a phospholipid bilayer], laced throughout with integral proteins. The lipid bilayer contains porins which allow the passage of molecules which are 10,000 Daltons or less. This permeability in the outer membrane allows for water, ions, and some proteins to flow freely into the inter membrane space.

Mitochondria as ATP Consumer

It is well documented mitochondria produce the ATP necessary for life, and that a proton gradient and membrane potential are required in order for ATP synthesis to occur in the mitochondria. However, mitochondria is very dynamic in that it can reverse its process. Complex V is the enzyme responsible for final ATP synthesis at the end of oxidative phosphorylation. When the trans-membrane potential is insufficient for ATP production, complex V, or F1F0-ATP synthase, can reverse its process, instead hydrolyzing ATP in order to pump protons out of the mitochondrial matrix in order to restore the proper gradient.

In a normal mitochondria performing respiration, ATP is removed through the adenine nucleotide translocase, or ANT. This helps maintain the trans-membrane potential, favoring phosphorylation of ADP. However, ATP hydrolysis is favored when the ANT is reverse, shuttling in ATP from glycolysis.

The consumption of ATP by mitochondria can clearly be potentially lethal to cells in conditions of extreme proton gradient degradation. It can also serve as a potential mechanism used by pathogens in the case of diseases related to mitochondrial respiration inhibition, (such as lack of oxygen in stroke or heart attack, or in less extreme cases where mitochondria are affected such as Alzheimer’s or Parkinson’s).

This reversal process of complex V is directly affected by IF1, the inhibitory factor of F1F0-ATPase. In response to acidification of the matrix of the mitochondria, the IF1 protein inhibits the activity of complex V as ATPase, usually in conjunction with the halting of respiration in conditions such as hypoxia or ischaemia. There is still much to learn about the mechanisms of IF1, however it is the major factor in protecting cells from ATP depletion in hypoxic conditions.

The crystal structure of IF1 is known. The protein acts as a homodimer, inhibiting two F1-ATPase units synchronously. There are many residues in the protein complex that form many associations with subunits of the F1-ATPase. It is believed that full association of IF1 with the F1F0-ATPase occurs only during ATP hydrolysis. It is suggested that IF1 may loosely bind to the F1-ATPase even during normal respiration, and that it may also aid in the efficiency of oxidative phosphorylation by serving as a ‘coupling factor.’

Mitochondria:ROS and Authophagy

The cite of Reactive Oxygen Species production with in the cell is found in the mitochondria. Reactive oxygen species (ROS) are small, highly reactive molecules that are short-lived. An incomplete one-electron reduction of oxygen is how ROS is formed.ROS includs oxygen anions, free radicals, and peroxides. Autophagy is one of the signaling pathways of the redox regulation of proteins by levels of ROS. Stress conditions activate autophagy, but pathological conditions deregulate autophagy. A build up of ROS can lead to oxidative stress that causes cellular constituents to be oxidized and damaged. Non-enzymatic and enzymatic antioxidizing agents have been created by the cell to prevent oxidative stress. Antioxidants are natural downregulators of ROS inducing autophagy. Autophagy is inhibited by TIGAR.

Endosymbiont Theory

In 1905, Mereschkowsky, a Russian biologist, published a paper on the theory that photosynthetic bacteria are the ancestors of modern day plant chloroplasts. Though this research was mostly ignored for several years, scientists came to see the similarities between isolated living bacteria and eukaryotic mitochondria. It is now largely accepted that mitochondria are descendants of "free-living" bacteria that were engulfed and incorporated as organelles by eukaryotic cells. The endosymbiont theory was further confirmed when mitochondria were discovered to contain their own DNA. It was confirmed even more so with the discovery that the mtDNA made enzymes and proteins that were needed for its own functionalities. The fact that the mitochondria also contains a double membrane also depicts the notion that it was originally a free living organism that was later ingested into another host. Mitochondria are produced by other mitochondria that act as "structural templates". A cell's two DNA genomes are still not aware of how mitochondrial membranes are assembled, showing that a mitochondria's structure isn't dictated by our DNA but must be passed on to future generations.

Mitochondria DNA

Mitochondria are the energy processing organelle that is found in the cell. Alongside with chloroplast, mitochondria are part endosymbiont theory, as stated above. The endosymbiont theory clearly stated the following about mitochondria and chloroplasts: it is enclosed by a double-membrane, it is about the same size as bacteria, it has its own circular DNA, its ribosomes are bacteria-like, and it has prokaryotic activities such as respiration and photosynthesis (mitochondria and chloroplasts respectively). Focusing on the third point about having its own circular DNA, mitochondria DNA (mtDNA) was found to be able to self-translate and replicate itself.

Genetics of mtDNA:

Unlike nuclear DNA which is inherited from both mother and father, the mammalian mtDNA is only inherited from mother. The mitochondria in mammalian sperm are destroyed in the fertilized oocyte. However, the replication pathway of mtDNA is very similar to nuclear DNA. Before replication, mtDNA becomes unwind by TWINKLE, which is a protein used to undo the double-stranded DNA. After the double-stranded is undone, it is replicated at one end with the help of mtDNA polymerase. mtDNA polymerase starts to form another double-stranded starting at 5' end of the mtDNA. Another protein called mitochondrial single-stranded binding (mtSSB) helps stabilize the unwound conformation and stimulates DNA synthesis by the polymerase holoenzyme.

Unfortunately, mtDNA replication displays no strict phase specificity as in nuclear DNA synthesis. Therefore, segregation of heteroplasmic mtDNA mutation can occur as a cell divides.

Mitochondrial transcription:

The transcription mechanism in mitochondria is likely similar to transcription in nucleus. However, there are some differences between RNA synthesis in mitochondria and in nucleus. The individual strands of the mtDNA molecules are denoted heavy strand (guanine rich) and light strand (guanine poor). This nucleotide bias explains why some codons are rare or absent in mitochondrial RNA.

The compact mammalian mtDNA genome lacks introns. The entire strand codes for either proteins, rRNA, or tRNA. Therefore, there is no need for slicing process in mitochondria

Each of the protein and rRNA genes is immediately flanked by at least one tRNA gene.

Some Interesting Discoveries about Mitochondria:

1. Albert Claude, who was a Belgian biochemist discovered in the first half of the last century discovered that Mitochondria catalyzed respiration. He did this by isolating them through centrifugation.

2. Scientists started from there and managed to map out the flow of electrons in cellular respiration in the past two decades.

3. Peter Mitchell then discovers that the key to the flow of free energy in respiration and photosynthesis is stored within the ion gradient across membranes. He receives a Nobel Prize for it in 1978.

4. Mitochondria is bordered by 2 membranes and, and holds one tenth of the cell’s proteins. Mitochondria also converts 10,000 to 50,000 times more energy per second than the sun does.

5. Mitochondria was also discovered to play a pivotal role in programmed cell death, or apoptosis. This shows mitochondria to thus also be part of the signal transduction network in the cell. For programmed cell death, Mitochondria first releases proteins called cytochromes into the cell’s cytoplasm. It is these signals that could potentially release proteases and nucleases onto the cell and trigger cellular suicide.

6. Isolated Mitochondria were discovered to produce their own proteins even though the identity of these proteins are yet to be determined.

7. It is a theory that Mitochondria evolved from bacteria, which explains why it is such an independent organelle and doesn’t seem to depend on other parts of the cell for survival. Another evidence for this rests in the fact that the mechanism for protein synthesis in mitochondria is similar to that in bacteria.

8. Mitochondria spread by growth and division of previously existing mitochondria. Mitochondria are thus able to tell building blocks for new mitochondria where to go and what to do.

9. Recent discoveries have revealed that mitochondria actually have a lot of extramitochondrial molecules that help regulate the expression of genes that turn into mitochondrial proteins. Peroxisomal-proliferator-actived receptor coactivator 1 (PGC1) plays a major role in this process.

10. The space in between the mitochondria’s membranes was recently discovered to be able to oxidize sulfhydryl groups to disulfide bridges even though that space is surrounded by highly reducing environments.

Role in Aging

Recent research into aging has lead scientists to believe that damage done by free radical oxygen may be one reason why organisms die from old age. Reactive Oxygen Species, or ROS, are produced in greatest quantity at the mitochondria, so this organelle is the most likely to be damaged by the free radical oxygen. One theory of aging says that damage done to the mitochondria by ROS harms the mtDNA, which reduces the operating capacity of the mitochondria. Mutations on the mtDNA could result in reduced ATP production, increased ROS production, and eventual apoptosis. The increased ROS production has the added affect of potentially being harmful to the cell hosting the mitochondria, which could cause mutation in the cell DNA.

Anti-aging research has shown in some model organisms that by genetically disrupting the function of mitochondria life span has been increased. This is because ROS production was decreased so that less damage was done to the mtDNA. Specifically, a reduction in the mitochondrial function of the electron transport chain (ETC) in c. elegans increased longevity of the organism.

Scientists hope to be able to apply this to extending human lifespan by somehow incorporating reduced mitochondrial function with a treatment of dietary restriction (DR). By reducing the amount of calories taken in, but not to the point of starvation, cellular processes would change such that emphasis is placed on maintaining existing cellular structures rather than generating new structures to replace old structures. This would cause cells to persist in the body longer and there would be fewer mutations due to gene replication during mitosis.

References

IF1: setting the pace of the F1Fo-ATP synthase. Campanella, M. Trends in Biochemical Sciences, Volume 34, Issue 7, 343-350, 25 June 2009.

Scherz-Shouval, Ruth, et al. Regulation of autophagy by ROS: physiology and pathology

Schatz, Gottfried. The Magic Garden, Annual Review of Biochemistry, 2007: 673-78.

Falkenberg, M., Larsson, N., & Gustafsson, C. M. (2007). DNA replication and transcription in mammalian mitochondria. Annual Review of Biochemistry, 679-699.

Mair, William, and Andrew Dillin. "Aging and Survival: The Genetics of Life Span Extension by Dietary Restriction." Annual Review of Biochemistry 77.1 (2008): 727-54. Web

Wager, Peter. Exercise Physiology. Aug 17,2011.