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== Abstract ==
== Abstract ==
Heart diseases are the leading cause of death for Americans today. Mitochondria play a crucial role in recovery following ischemia (blood flow restriction) and reperfusion (blood flow return) injury, when a surge of reactive oxygen species (radicals) originating from the mitochondrial electron transport chain causes damage to proteins, lipids and DNA. Uncoupling protein 2 (UCP2), an inner mitochondrial membrane transport protein, is speculated to participate in this protection. The presumed function of UCP2 is carrying protons (H+) into the mitochondrial matrix along a concentration gradient generated by the electron transport chain. Normally, this proton (H+) gradient is used by ATP synthase to phosphorylate ADP to ATP. Under certain conditions, protons (H+) may preferentially be transported through UCP2, creating a detour past ATP synthase (“uncoupling”). Such uncoupling reduces damaging reactive oxygen species whose presence may actually activate UCP2 by residue modification. There are two proposed mechanisms for the transport of protons (H+) into the matrix. One is the direct transport of protons (H+) through UCP2. Alternatively, a fatty acid anion is transported out of the matrix through UCP2, while the protonated fatty acid permeates through the membrane into the matrix. UCP2 must be tightly regulated so it is only active when required, enabling the mitochondria to produce ATP. Understanding transport mechanism and regulation of UCP2 could lead to effective prevention of tissue injury due to heart attack. The Brookfield Central High School SMART Team created a physical model of UCP2 using 3-D modeling printing technology in order to better understand the structure-function relationship of UCP2.
Heart diseases are the leading cause of death for Americans today. Mitochondria play a crucial role in recovery following ischemia (blood flow restriction) and reperfusion (blood flow return) injury, when a surge of reactive oxygen species (radicals) originating from the mitochondrial electron transport chain causes damage to proteins, lipids and DNA. Uncoupling protein 2 (UCP2), an inner mitochondrial membrane transport protein, is speculated to participate in this protection. The presumed function of UCP2 is carrying protons (H+) into the mitochondrial matrix along a concentration gradient generated by the electron transport chain.<ref name="Berardi">PMID:21785437</ref> Normally, this proton (H+) gradient is used by ATP synthase to phosphorylate ADP to ATP. Under certain conditions, protons (H+) may preferentially be transported through UCP2, creating a detour past ATP synthase (“uncoupling”). Such uncoupling reduces damaging reactive oxygen species whose presence may actually activate UCP2 by residue modification.<ref name="Krauss">PMID:15738989</ref> There are two proposed mechanisms for the transport of protons (H+) into the matrix. One is the direct transport of protons (H+) through UCP2. Alternatively, a fatty acid anion is transported out of the matrix through UCP2, while the protonated fatty acid permeates through the membrane into the matrix. UCP2 must be tightly regulated so it is only active when required, enabling the mitochondria to produce ATP. Understanding transport mechanism and regulation of UCP2 could lead to effective prevention of tissue injury due to heart attack. The Brookfield Central High School SMART Team created a physical model of UCP2 using 3-D modeling printing technology in order to better understand the structure-function relationship of UCP2.
== Data Piece ==
[[Image:BCSMART_11-12_Figure_7.JPG|center|375px]]
'''The Effect of UCP2 on ROS Production'''
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The data above .<ref name="Teshima">PMID: 12855674</ref> shows the effect of UCP2 on ROS production. The ROS is made visible by DCF fluorescence(dichloro-fluorescein, which fluoresces in the presence of H2O2). ROS species are introduced into neonatal rat cardiac ventricular myocytes by injecting H2O2.
<br>
A. Confocal microscope images of cardiomyocytes’ ROS production
 
Top row: ROS in myoctes which have been exposed to adenovirus without the UCP2 protein. Increased DCF fluorescence can be seen over time, representing the presence of a significant level of ROS.     
 
Bottom row: ROS in a myocytes exposed to an adenovirus producing the UCP2 protein. DCF fluorescence is barely visible, representing little to no ROS presence.
 
 
B. Quantitative representation of the changes in ROS levels corresponding to the fluorescence images in A using time-lapse confocal microscopy. The presence of UCP2 greatly reduces ROS levels, limiting damage to cardiac ventricular myocytes.
== Figures ==
== Figures ==
[[Image:BCSMART_11-12_Figure_1.JPG|left|375px|thumb|'''FIGURE 1: Movement of Protons (H+) Across Inner Mitochondrial Membrane'''<br>
[[Image:BCSMART_11-12_Figure_1.JPG|left|375px|thumb]]
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'''FIGURE 1: Movement of Protons (H+) Across Inner Mitochondrial Membrane'''
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1. The electron transport chain pumps protons (H+) into the inter membrane space.
1. The electron transport chain pumps protons (H+) into the inter membrane space.
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of ADP to ATP.
of ADP to ATP.
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]]
[[Image:BCSMART_11-12_Figure_2.JPG|right|375px|thumb|'''FIGURE 2: ROS Production'''
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[[Image:BCSMART_11-12_Figure_2.JPG|left|375px|thumb]]
 
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'''FIGURE 2: ROS Production'''
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1. During reperfusion (return of blood flow), the electron transport chain sends more protons (H+) into the inter membrane space, producing a high concentration.
1. During reperfusion (return of blood flow), the electron transport chain sends more protons (H+) into the inter membrane space, producing a high concentration.
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that carry a negative charge (O2
that carry a negative charge (O2
-)]]
-)]]
[[Image:BCSMART_11-12_Figure_3.JPG|left|375px|thumb|'''FIGURE 3: UCP2 Relieving the High Concentration Gradient'''
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[[Image:BCSMART_11-12_Figure_3.JPG|left|375px|thumb]]
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'''FIGURE 3: UCP2 Relieving the High Concentration Gradient'''
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One way the mitochondria can alleviate the proton buildup in the inter membrane space is to transport the protons (H+) back into the matrix. One possible protein involved in this transportation is uncoupling protein 2 (UCP2).
One way the mitochondria can alleviate the proton buildup in the inter membrane space is to transport the protons (H+) back into the matrix. One possible protein involved in this transportation is uncoupling protein 2 (UCP2).
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1.UCP2 brings protons (H+) into the matrix, relieving the high concentration gradient.
1.UCP2 brings protons (H+) into the matrix, relieving the high concentration gradient.
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2.The “back up” of electrons and radical production is reduced, minimizing damage to cardiomyocytes.
2.The “back up” of electrons and radical production is reduced, minimizing damage to cardiomyocytes.
]]
]]
[[Image:BCSMART_11-12_Figure_4.JPG|right|375px|thumb|'''FIGURE 4: Proposed Mechanisms for UCP2 Transport of Protons (H+)'''
[[Image:BCSMART_11-12_Figure_4.JPG|left|375px|thumb]]
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There are two proposed mechanisms for UCP2 transport of protons (H+) into the mitochondria.
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1. Direct proton (H+) transport via negatively charged UCP2 residues through the membrane.
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2. A neutral fatty acid is flipped through the mitochondrial membrane into the matrix, and because of the basic environment, it releases the proton (H+). The now negatively charged anion is then transported
through the membrane by UCP2.]]
[[Image:BCSMART_11-12_Figure_5.JPG|left|375px|thumb|'''FIGURE 5: UCP2 Protein Structure'''
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UCP2 is an inner mitochondrial membrane protein consisting of 6 transmembrane helices, shown in
orange, with both the N- and C- termini pointing to the inter membrane space. UCP2 activity is inhibited
by nucleotide binding (eg. GDP). Lys141 and Arg185, which are speculated to bind to GDP, are
shown in light sky blue.]]
[[Image:BCSMART_11-12_Figure_6.JPG|right|375px|thumb|'''FIGURE 6: View of UCP2 Down the Barrel'''
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View from matrix side through the putative transport path of UCP2. Our model may support the mechanism
where a protonated fatty acid flips through the inner membrane into the matrix, loses its proton
due to the basic interior environment, and is attracted to the matrix side by positively charged residues
(navy). Additional positively charged residues in the interior of the barrel may pass the fatty acid anion
towards the inter membrane space.]]
[[Image:BCSMART_11-12_Figure_7.JPG|middle|375px|thumb|'''FIGURE 7: The Effect of UCP2 on ROS Production'''
<br>
<br>
The data above shows the effect of UCP2 on ROS production. The ROS is made visible by DCF fluorescence(dichloro-fluorescein, which fluoresces in the presence of H2O2). ROS species are introduced into neonatal rat cardiac ventricular myocytes by injecting H2O2.
 
'''FIGURE 4: Proposed Mechanisms for UCP2 Transport of Protons (H+)'''
<br>
<br>
A. Confocal microscope images of cardiomyocytes’ ROS production
There are two proposed mechanisms for UCP2 transport of protons (H+) into the mitochondria.
Top row: ROS in myoctes which have been exposed to adenovirus without the UCP2 protein. Increased DCF fluorescence can be seen over time, representing the presence of a significant level of ROS. Bottom row: ROS in a myocytes exposed to an adenovirus producing the UCP2 protein.DCF fluorescence is barely visible, representing little to no ROS presence.
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<br>
B. Quantitative representation of the changes in ROS levels corresponding to the fluorescence images in A using time-lapse confocal microscopy.
1. Direct proton (H+) transport via negatively charged UCP2 residues through the membrane.
<br>
<br>
The presence of UCP2 greatly reduces ROS levels, limiting damage to cardiac ventricular myocytes.]]
2. A neutral fatty acid is flipped through the mitochondrial membrane into the matrix, and because of the basic environment, it releases the proton (H+). The now negatively charged anion is then transported
through the membrane by UCP2.]]
 
<StructureSection load='2lck_pdb' scene='Sandbox_Reserved_508/2lck_pdb/12' size='750' side='middle' caption='Uncoupling Protein 2, 2lck <ref name="Berardi">PMID:21785437</ref>Structure of UCP 2 displaying key features. In this model of UCP2, the light sea green parts represent the alpha helices which are thought to be involved in the transport of protons. All negatively-charged amino acids, glutamic and aspartic, are colored crimson. These may be involved in the direct transport of protons through UCP2. All positively charged amino acids, except for Lys141 and Arg185, are colored navy. These may be involved in the transport of deprotonated fatty acid molecules through UCP2, as well. Lys141 and Arg185, which are speculated to bind to the UCP2 inhibitor, GDP, are colored light sky blue. Helix sequences, conserved between different UCP proteins, which may also have a role in fatty acid transport, are colored orange. Helices found in the matrix or the inter-membrane space are colored light aquamarine. Loops, which are not involved in the active site, are colored white.  '>
 
 
==References==
<references />
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