Sandbox Reserved 641: Difference between revisions

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Glutamate dehydrogenase (GDH)is an enzyme found in the mitochondria of most organisms. GDH is used to remove the ketone group and replace it with an α-amine group on the α-carbon, which forms glutamate. Glutamate is one of the 20 essential amino acids. This is done in reverse to supply α-ketoglutarate to the tricarboxylic acid (TCA) cycle. GDH is an oxidoreductase, which is an enzyme that transfers electrons from one molecule (reductant/electron donor) to another molecule (oxidant/electron acceptor).

Glutamate dehydrogenase is a hexamer that contains two domains that have three subunits. GHD contains approximately 18 alpha helices and thirteen beta sheets. There is a large cleft that separates the two domains and allows for a substrate to enter and bind. The protein then closes around the substrate. For mammals only there is a structure that extends outward of the protein called the "antennae."

Glutamate dehydrogenase is a hexamer that contains two domains that have three subunits. GHD contains approximately 18 alpha helices and thirteen beta sheets. There is a large cleft that separates the two domains and allows for a substrate to enter and bind. The protein then closes around the substrate. For mammals only there is a structure that extends outward of the protein called the "antennae." (8)

Today, one of the primary research uses for glutamate dehydrogenase is to determine how well the human liver is functioning. If the level of GDH is too high that could indicate necrosis of the liver.

Today, one of the primary research uses for glutamate dehydrogenase is to determine how well the human liver is functioning. If the level of GDH is too high that could indicate necrosis of the liver. (7)

== '''Structure''' ==

Glutamate Dehydrogenase is a hexamer that is comprised of two trimer subunits. These two subunits are stacked on top of each other and composed of three domains. The top of each domain contains a "NAD-binding domain" that has the conserved nucleotide-binding motif. A larger helix-loop-helix structure rises above this and is referred to as an "antenna." This antenna contains approximately 50 amino acids and is thought to play a major role in regulation of the enzyme. This antennae structure is only found in animals. (1) The bottom domain contacts a domain in the other trimer, holding the two trimers together. The total size of each of the subunits is approximately 56.1 kD and 506 amino acids long. (5)

When a substrate binds to the enzyme it binds to the deep recess of the cleft between the NAD binding domain and the lower domain. Along the outside surface of the cleft a coenzyme (NAD+) binds to the C-terminal end causing the binding domain to rotate by about 18 degrees and close down on the substrate and coenzyme. The active sites for GDH are located at residues around 182-187. <scene name='Sandbox_Reserved_641/Active_sites/1'>Active Sites</scene> Residues at the locations of 200-206, 375-370, and 421-423 are involved in closing the cleft between the domains. As the cleft is closing the antenna pushes against the pivot helix of the adjacent subunit. The pivot helix rotates counter clockwise around both the helical axis and the trimer 3-fold axis. The hexamer then compresses the inner core showing that catalysis involves the entire hexamer. (8)

When a substrate binds to the enzyme it binds to the deep recess of the cleft between the NAD binding domain and the lower domain. Along the outside surface of the cleft a coenzyme (NAD+) binds to the C-terminal end causing the binding domain to rotate by about 18 degrees and close down on the substrate and coenzyme. (2) The active sites for GDH are located at residues around 182-187. <scene name='Sandbox_Reserved_641/Active_sites/1'>Active Sites</scene> Residues at the locations of 200-206, 375-370, and 421-423 are involved in closing the cleft between the domains. As the cleft is closing the antenna pushes against the pivot helix of the adjacent subunit. The pivot helix rotates counter clockwise around both the helical axis and the trimer 3-fold axis. The hexamer then compresses the inner core showing that catalysis involves the entire hexamer. (8)

The figure on the right shows the two domains of GDH. The orange represent the glutamate binding sites and the blue sites represent the domain involved in assembly of the hexamer.

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As NADH is oxidized, the decrease in the absorbance per minute is measured spectrophotometrically at 340nm and is proportional to the GLDH activity.

An ~~Example~~ of this can be seen here: [https://docs.google.com/a/ncsu.edu/viewer?a=v&pid=gmail&attid=0.2&thid=13addc1ac3f18236&mt=application/pdf&url=https://mail.google.com/mail/u/0/?ui%3D2%26ik%3D5ad9b5051c%26view%3Datt%26th%3D13addc1ac3f18236%26attid%3D0.2%26disp%3Dsafe%26realattid%3Df_h998el621%26zw&sig=AHIEtbSNiAIgz_br1UY0dW534zXhAWFV4A]

An example of this can be seen here: [https://docs.google.com/a/ncsu.edu/viewer?a=v&pid=gmail&attid=0.2&thid=13addc1ac3f18236&mt=application/pdf&url=https://mail.google.com/mail/u/0/?ui%3D2%26ik%3D5ad9b5051c%26view%3Datt%26th%3D13addc1ac3f18236%26attid%3D0.2%26disp%3Dsafe%26realattid%3Df_h998el621%26zw&sig=AHIEtbSNiAIgz_br1UY0dW534zXhAWFV4A]

== Notes ==

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 <Structure load='1V9L' size='500' frame='true' align='right' caption='Glutamate Dehydrogenase' scene='Insert optional scene name here' />
 Glutamate dehydrogenase (GDH)is an enzyme found in the mitochondria of most organisms.  GDH is used to remove the ketone group and replace it with an α-amine group on the α-carbon, which forms glutamate.  Glutamate is one of the 20 essential amino acids.  This is done in reverse to supply α-ketoglutarate to the tricarboxylic acid (TCA) cycle.  GDH is an oxidoreductase, which is an enzyme that transfers electrons from one molecule (reductant/electron donor) to another molecule (oxidant/electron acceptor).
-Glutamate dehydrogenase is a hexamer that contains two domains that have three subunits.  GHD contains approximately 18 alpha helices and thirteen beta sheets.  There is a large cleft that separates the two domains and allows for a substrate to enter and bind.  The protein then closes around the substrate.  For mammals only there is a structure that extends outward of the protein called the "antennae."
+Glutamate dehydrogenase is a hexamer that contains two domains that have three subunits.  GHD contains approximately 18 alpha helices and thirteen beta sheets.  There is a large cleft that separates the two domains and allows for a substrate to enter and bind.  The protein then closes around the substrate.  For mammals only there is a structure that extends outward of the protein called the "antennae." (8)
-Today, one of the primary research uses for glutamate dehydrogenase is to determine how well the human liver is functioning.  If the level of GDH is too high that could indicate necrosis of the liver.
+Today, one of the primary research uses for glutamate dehydrogenase is to determine how well the human liver is functioning.  If the level of GDH is too high that could indicate necrosis of the liver. (7)
 == '''Structure''' ==
 Glutamate Dehydrogenase is a hexamer that is comprised of two trimer subunits.  These two subunits are stacked on top of each other and composed of three domains.  The top of each domain contains a "NAD-binding domain" that has the conserved nucleotide-binding motif.  A larger helix-loop-helix structure rises above this and is referred to as an "antenna."  This antenna contains approximately 50 amino acids and is thought to play a major role in regulation of the enzyme.  This antennae structure is only found in animals. (1)  The bottom domain contacts a domain in the other trimer, holding the two trimers together.  The total size of each of the subunits is approximately 56.1 kD and 506 amino acids long. (5)
-When a substrate binds to the enzyme it binds to the deep recess of the cleft between the NAD binding domain and the lower domain.  Along the outside surface of the cleft a coenzyme (NAD+) binds to the C-terminal end causing the binding domain to rotate by about 18 degrees and close down on the substrate and coenzyme.  The active sites for GDH are located at residues around 182-187.  <scene name='Sandbox_Reserved_641/Active_sites/1'>Active Sites</scene>  Residues at the locations of 200-206, 375-370, and 421-423 are involved in closing the cleft between the domains.   As the cleft is closing the antenna pushes against the pivot helix of the adjacent subunit.  The pivot helix rotates counter clockwise around both the helical axis and the trimer 3-fold axis.  The hexamer then compresses the inner core showing that catalysis involves the entire hexamer. (8)
+When a substrate binds to the enzyme it binds to the deep recess of the cleft between the NAD binding domain and the lower domain.  Along the outside surface of the cleft a coenzyme (NAD+) binds to the C-terminal end causing the binding domain to rotate by about 18 degrees and close down on the substrate and coenzyme. (2)  The active sites for GDH are located at residues around 182-187.  <scene name='Sandbox_Reserved_641/Active_sites/1'>Active Sites</scene>  Residues at the locations of 200-206, 375-370, and 421-423 are involved in closing the cleft between the domains.   As the cleft is closing the antenna pushes against the pivot helix of the adjacent subunit.  The pivot helix rotates counter clockwise around both the helical axis and the trimer 3-fold axis.  The hexamer then compresses the inner core showing that catalysis involves the entire hexamer. (8)
 The figure on the right shows the two domains of GDH.  The orange represent the glutamate binding sites and the blue sites represent the domain involved in assembly of the hexamer.
@@ Line 42: / Line 42: @@
 As NADH is oxidized, the decrease in the absorbance per minute is measured spectrophotometrically at 340nm and is proportional to the GLDH activity.
-An Example of this can be seen here: [https://docs.google.com/a/ncsu.edu/viewer?a=v&pid=gmail&attid=0.2&thid=13addc1ac3f18236&mt=application/pdf&url=https://mail.google.com/mail/u/0/?ui%3D2%26ik%3D5ad9b5051c%26view%3Datt%26th%3D13addc1ac3f18236%26attid%3D0.2%26disp%3Dsafe%26realattid%3Df_h998el621%26zw&sig=AHIEtbSNiAIgz_br1UY0dW534zXhAWFV4A]
+An example of this can be seen here: [https://docs.google.com/a/ncsu.edu/viewer?a=v&pid=gmail&attid=0.2&thid=13addc1ac3f18236&mt=application/pdf&url=https://mail.google.com/mail/u/0/?ui%3D2%26ik%3D5ad9b5051c%26view%3Datt%26th%3D13addc1ac3f18236%26attid%3D0.2%26disp%3Dsafe%26realattid%3Df_h998el621%26zw&sig=AHIEtbSNiAIgz_br1UY0dW534zXhAWFV4A]
 == Notes ==