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hexokinase is an enzyme that phosphorylates a six-carbon sugar, a hexose, to a hexose phosphate. In most tissues and organisms, glucose is the most important substrate of hexokinases, and glucose 6-phosphate the most important product. Hexokinases have been found in every organism checked, ranging from bacteria, yeast, and plants, to humans and other vertebrates. They are categorized as actin fold proteins, sharing a common ATP binding site core surrounded by more variable sequences that determine substrate affinities and other properties. Several hexokinase isoforms or isozymes providing different functions can occur in a single species.  
== '''Hexokinase''' ==
 
 
Hexokinase is an enzyme that phosphorylates a six-carbon sugar, a hexose, to a hexose phosphate. In most tissues and organisms, glucose is the most important substrate of hexokinases, and glucose 6-phosphate the most important product. Hexokinases have been found in every organism checked, ranging from bacteria, yeast, and plants, to humans and other vertebrates. They are categorized as actin fold proteins, sharing a common ATP binding site core surrounded by more variable sequences that determine substrate affinities and other properties. Several hexokinase isoforms or isozymes providing different functions can occur in a single species.  


-Hexokinase I/A is found in all mammalian tissues, and is considered a "housekeeping enzyme," unaffected by most physiological, hormonal, and metabolic changes.  
-Hexokinase I/A is found in all mammalian tissues, and is considered a "housekeeping enzyme," unaffected by most physiological, hormonal, and metabolic changes.  
Line 7: Line 10:
-Hexokinase III/C is substrate-inhibited by glucose at physiologic concentrations. Little is known about the regulatory characteristics of this isoform.  
-Hexokinase III/C is substrate-inhibited by glucose at physiologic concentrations. Little is known about the regulatory characteristics of this isoform.  


-Hexokinase IV/D is also known as glucokinase and is described below.  
-Hexokinase IV/D is also known as glucokinase.
 
{{STRUCTURE_1hkc |  PDB=1hkc  |  SCENE=  }}
 
== Structure of Hexokinase ==
Hexokinase is composed of an N-terminal regulatory domain and a C-terminal catalytic domain. These two domains are <scene name='Bawel_sandbox1/Hexokinase/2'>joined together by an alpha helix</scene>. The molecular weights of hexokinases are around 100 kD. Each domain weighs about 50kD and contains a <scene name='Bawel_sandbox1/Glucose_binding_site/1'>glucose binding site</scene>. But, only in hexokinase II do both halves have functional active sites. The tertiary structure of hexokinase includes an open alpha/beta sheet. There is a large amount of variation associated with this structure. The ATP-binding domain is composed of <scene name='Bawel_sandbox1/5_beta_sheets/3'>five beta sheets and two alpha helices</scene>. In this open alph/beta sheet four of the beta sheets are parallel and one is in the anitparallel directions. The alpha helices and beta loops connect the beta sheets to produce this open alpha/beta sheet.




[[Image:Hexokinase_mechanism2.GIF|350px|left|thumb]]


== Mechanism of Hexokinase ==
In the first reaction of glycolysis, the gamma-phosphoryl group of an ATP molecule is transferred to the oxygen at the C-6 of glucose. Hexokinase catalyzes this phosphoryl group transfer. To start this reaction, ATP forms a complex with magnesium (II) ion and glucose binds to hexokinase. The magnesium-ATP complex then binds with the hexokinase-glucose complex and forms an intermediate (Zeng, et al. present a picture showing the interctions of brain hexokinase with ATP). <scene name='Bawel_sandbox1/Asp_532_and_thr_680/2'>Asp 532 and Thr 680</scene> are thought to be involved in binding the magnesium ion in the magnesium-ATP complex [4]. The hydroxyl group on the terminal phosphoryl group of the ATP molecule nucleophilically attacks carbon 6 on glucose. This produces glucose-6-phosphate still bound to hexokinase and ADP still in complex with magnesium ion [5]. Glucose-6-phosphate and the magnesium-ADP complex leave hexokinase. Glucose-6-phosphate and ADP are the products of this reaction. Hexokinase undergoes an induced-fit conformational change when it binds to glucose, which ultimately prevents the hydrolysis of ATP. It also experiences potent allosteric inhibition under physiological concentrations by its immediate products, glucose-6-phosphate [4]. This is a mechanism by which the influx of substrate into the glycolytic pathway is controlled.


Hexokinase Structure:
The tertiary structure of hexokinase includes an open alpha/beta sheet. There is a large amount of variation associated with this structure. The ATP-binding domain is composed of five beta sheets and three alpha helices. In this open alph/beta sheet four of the beta sheets are parallel and one is in the anitparallel directions. The alpha helices and beta loops connect the beta sheets to produce this open alpha/beta sheet. The crevice indicates the ATP-binding domain of this glycolytic enzyme. The molecular weights of hexokinases are around 100 kD. Each consists of two similar 50kD halves, but only in hexokinase II do both halves have functional active sites.


Mechanism of Hexokinase:
== Kinetics and Inhibition of Hexokinase ==
In the first reaction of glycolysis, the gamma-phosphoryl group of an ATP molecule is transferred to the oxygen at the C-6 of glucose. Hexokinase catalyzes this phosphoryl group transfer. To start this reaction, ATP forms a complex with magnesium (II) ion and glucose binds to kexokinase. The magnesium-ATP complex then binds with the hexokinase-glucose complex and forms an intermediate (Zeng, et al. present a picture showing the interctions of brain hexokinase with ATP). The hydroxyl group on the terminal phosphoryl group of the ATP molecule nucleophilically attacks the carbon 6 on glucose. This produces glucose-6-phosphate still bound to hexokinase and ADP still in complex with magnesium ion [5]. Glucose-6-phosphate and the magnesium-ADP complex leave hexokinase. Glucose-6-phosphate and ADP are the products of this reaction. Hexokinase undergoes an induced-fit conformational change when it binds to glucose, which ultimately prevents the hydrolysis of ATP. It also experiences potent allosteric inhibition under physiological concentrations by its immediate products, glucose-6-phosphate [4]. This is a mechanism by which the influx of substrate into the glycolytic pathway is controlled.


Hexokinase activates glycoloysis by phosphorylating glucose. Since the phosphorylation of glucose to glucose-6-phosphate is the rate limiting step of glucose metabolism, hexokinase has a very important role in regulating healthy glucose levels in the human body [7]. Hexokinase has high affinity, thus a low Km, for glucose. Tissues where hexokinase is present use glucose at low blood serum levels.


G6P inhibits hexokinase by binding to the N-terminal domain(this is simple feedback inhibition). It competitively inhibits the binding of ATP [8]. If the cell is not using the G6P that it is making, then it stops making it. In this way, hexokinase can also slow down glycolysis. Hexokinase I is thought to be the "pacemaker" of glycolysis in brain tissue and red blood cells [4]. Inorganic phosphate allosterically relieves hexokinase of inhibition by G6P [8].


[[Image:hexokinase-glucose.png|350px|left|thumb]]






[edit] Glucokinase, an Isoenzyme of Hexokinase: Glucokinase (hexokinase D) is a monomeric cytoplasmic enzyme. As an isoenzyme of hexokinase, glucokinase is capable of prompting the first step of glycogen synthesis and glycolysis: the phosphorylation of glucose to glucose-6-phosphate (G6P). Since the phosphorylation of glucose to glucose-6-phosphate is the rate limiting step of glucose metabolism, glucokinase has a very important job in regulating healthy glucose levels in the human body [7].Mainly found in the liver and pancreas, glucokinase can also be found in the gut and brain.


Glucokinase vs. Other Hexokinases: Glucokinase is unique from other hexokinase in its kinetic properties and it is coded by a different gene. Compared to other hexokinases, glucokinase has a lower affinity, thus a higher Km, for glucose. Glucokinase actively stores glucose when serum glucose levels are high. Tissues lacking glucokinase use glucose at lower serum levels, so they use hexokinase with higher affinity for glucose. Also, G6P inhibits hexokinase (this is simple feedback inhibition). If the cell is not using up the G6P that it is making, then it should stop making it. However, G6P does not inhibit glucokinase. In fact, the steady state velocity of glucose phosphorylation displays a sigmoidal response curve to increasing glucose concentrations [7]. Glucokinase responds quickly to changes in glucose concentration becuase of these unique kinetic proeprties.


   
   
Structure of Glucokinase


Glucokinase Structure: Glucokinase also contains inactive and active conformations. Glucokinase consists of one chain or subunit of 448 amino acids forming a monomeric molecule consisting of 13 alpha helices and 5 beta sheets that can phosporylate glucose and other hexoses. The chain is folded into two distinct regions, a small and large domain. Glucokinase has one active binding site for glucose and one for ATP, which is the energy source for phosphorylation. This active binding site is located between the small and large domains. The carboxyl terminus is part of the alpha 13 helix, which codes for the region that forms half of the binding site for glucose. Glucokinase can be modulated to form an inactive and active complex. The inactive conformation forms when the alpha 13 helix has been modulated away from the rest of the molecule forming a large space. This space is too large to bind glucose so it is said to be in the inactive form. The alternative is when the alpha 13 helix is modulated to form a smaller space thus activating the protein[2].




Glucose forms hydrogen bonds at the bottom of the deep crevice between the large domain and the small domain at the glucose binding site (active form). E256, E290 (shown in green) of the large domain, T168, K169 (shown in red) of the small domain, and N204, D205 (shown in yellow) of a connecting region form hydrogen bonds with glucose. The glucose binding site (inactive form) has a different conformation. At the allosteric site (active form), ATP forms hydrogen bonds with R63 and Y215 (shown in orange) and hydrophobically interacts with M210, Y214 (shown in blue) of the α5 helix and V452, V455 (shown in green) of the α13 helix. The allosteric site (inactive form) again shows structural differences. The differences in these two conformations allows glucokinase to function properly in different levels of glucose concentration.
{{STRUCTURE_3cin |  PDB=3cin  |  SCENE=  }}




Role in Organ Systems: In the liver glucokinase increases the synthesis of glycogen and is the first step in glycolysis, the main producer of ATP in the body. Glucokinase is responsible for phospohorylating the majority of glucose in the liver and pancreas. Glucokinase only binds to and phosphorylates glucose when levels are higher than normal blood glucose level, allowing it to maintain constant glucose levels[2]. By phosphorylating glucose, glucokinase creates glucose 6-phosphate. Glucose 6-phosphate can then be used by the liver through the glycolytic pathway. Along with this process in the liver, glucokinase also facilitates glycogen synthesis. Through this the majority of the body's glucose is stored. Glucose 6-phosphate is also one of the starting materials of the TCA cycle which is responsible for the majority of ATP production in the body.


In the pancreas, a rise in glucose levels increases the activity of glucokinase causing an increase in glucose 6-phosphate. This causes the triggering of the beta cells to secret insulin[3]. Glucokinase is the first step in this reaction. Insulin then allows other cells in the body to take up glucose, actively lowering the glucose level. Around 200 mutations inactivating glucokinase have been identified in triggering diabetes mellitus and 5 mutations activating glucokinase have been identified in triggering hypoglycemia [6]. Clearly, glucose plays a crucial role in maintaining healthy sugar levels in humans. Studies looking at the structure of gluockinase reveal the presence of allosteric site. This site is only present after glucose has attached to glucokinase [6]. Certain small molecules bind to this allosteric site and increase the activity of glucokinase [6]. These small molecules can potentially be used as a treatment for diabetes. 


Hexokinase I is thought to be the "pacemaker of glycolysis in brain tissue and red blood cells [4].
 
 
 


[edit] Additional Resources
[edit] Additional Resources
Line 50: Line 54:


[edit] References
[edit] References
1.↑ Pollard-Knight D, Cornish-Bowden A. Mechanism of liver glucokinase. Mol Cell Biochem. 1982 Apr 30;44(2):71-80. PMID:7048063
1.↑ Pollard-Knight D, Cornish-Bowden A. Mechanism of liver glucokinase. Mol Cell Biochem. 1982 Apr 30;44(2):71-80. PMID:[[7048063]]
2.↑ 2.0 2.1 Kamata K, Mitsuya M, Nishimura T, Eiki J, Nagata Y. Structural basis for allosteric regulation of the monomeric allosteric enzyme human glucokinase. Structure. 2004 Mar;12(3):429-38. PMID:15016359 doi:10.1016/j.str.2004.02.005
 
3.↑ Postic C, Shiota M, Magnuson MA. Cell-specific roles of glucokinase in glucose homeostasis. Recent Prog Horm Res. 2001;56:195-217. PMID:11237213
2.↑ 2.0 2.1 Kamata K, Mitsuya M, Nishimura T, Eiki J, Nagata Y. Structural basis for allosteric regulation of the monomeric allosteric enzyme human glucokinase. Structure. 2004 Mar;12(3):429-38. PMID:[[15016359]] doi:10.1016/j.str.2004.02.005
 
3.↑ Postic C, Shiota M, Magnuson MA. Cell-specific roles of glucokinase in glucose homeostasis. Recent Prog Horm Res. 2001;56:195-217. PMID:[[11237213]]
 
4.↑ Zeng C, Aleshin A, Hardie J, Harrison R, Fromm H. ATP-Binding site of Human Brain Hexokinase as Studied by Molecular Modeling and Site-Directed Mutagenesis. Biochem. 1996 Aug 6;35:13157-13164.
4.↑ Zeng C, Aleshin A, Hardie J, Harrison R, Fromm H. ATP-Binding site of Human Brain Hexokinase as Studied by Molecular Modeling and Site-Directed Mutagenesis. Biochem. 1996 Aug 6;35:13157-13164.
5.↑ hammes G, and Kochavi D. Studies of the Enzyme Hexokinase: Kinetic Inhibition by Products. Massachusetts Institute of Technology. 1961 Oct 5.  
 
5.↑ hammes G, and Kochavi D. Studies of the Enzyme Hexokinase: Kinetic Inhibition by Products. Massachusetts Institute of Technology. 1961 Oct 5.
6.↑ Ralph E, Thomson J, Almaden J, Sun S. Glucose Modulation fo Glucokinase Activation by Small Molecules.  Biochem. 2008 Feb 15;47:5028-5036.
6.↑ Ralph E, Thomson J, Almaden J, Sun S. Glucose Modulation fo Glucokinase Activation by Small Molecules.  Biochem. 2008 Feb 15;47:5028-5036.
7.↑ Pal P, and Miller B. Activating Mutations in the Human Glucokinase Gene Revealed by Genetic Selection. Biochem. 2008 Dec 3;48:814-816.
7.↑ Pal P, and Miller B. Activating Mutations in the Human Glucokinase Gene Revealed by Genetic Selection. Biochem. 2008 Dec 3;48:814-816.
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8.↑ Aleshin A, Malfois M, Liu X, Kim C, Fromm H, Honzatko R, Koch M, Svergun D. Nonaggregating Mutant of Recombinant Human Hexokinase I Exhibits Wild-Type Kinetics and Rod-like Conformations in Solution. Biochem. 1999 Apr 29;38:8359-8366.
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Latest revision as of 22:23, 8 June 2011

HexokinaseHexokinase

Hexokinase is an enzyme that phosphorylates a six-carbon sugar, a hexose, to a hexose phosphate. In most tissues and organisms, glucose is the most important substrate of hexokinases, and glucose 6-phosphate the most important product. Hexokinases have been found in every organism checked, ranging from bacteria, yeast, and plants, to humans and other vertebrates. They are categorized as actin fold proteins, sharing a common ATP binding site core surrounded by more variable sequences that determine substrate affinities and other properties. Several hexokinase isoforms or isozymes providing different functions can occur in a single species.

-Hexokinase I/A is found in all mammalian tissues, and is considered a "housekeeping enzyme," unaffected by most physiological, hormonal, and metabolic changes.

-Hexokinase II/B constitutes the principal regulated isoform in many cell types and is increased in many cancers.

-Hexokinase III/C is substrate-inhibited by glucose at physiologic concentrations. Little is known about the regulatory characteristics of this isoform.

-Hexokinase IV/D is also known as glucokinase.


PDB ID 1hkc

Drag the structure with the mouse to rotate
1hkc, resolution 2.80Å ()
Ligands: , ,
Activity: Hexokinase, with EC number 2.7.1.1
Resources: FirstGlance, OCA, RCSB, PDBsum
Coordinates: save as pdb, mmCIF, xml



Structure of HexokinaseStructure of Hexokinase

Hexokinase is composed of an N-terminal regulatory domain and a C-terminal catalytic domain. These two domains are . The molecular weights of hexokinases are around 100 kD. Each domain weighs about 50kD and contains a . But, only in hexokinase II do both halves have functional active sites. The tertiary structure of hexokinase includes an open alpha/beta sheet. There is a large amount of variation associated with this structure. The ATP-binding domain is composed of . In this open alph/beta sheet four of the beta sheets are parallel and one is in the anitparallel directions. The alpha helices and beta loops connect the beta sheets to produce this open alpha/beta sheet.


Mechanism of HexokinaseMechanism of Hexokinase

In the first reaction of glycolysis, the gamma-phosphoryl group of an ATP molecule is transferred to the oxygen at the C-6 of glucose. Hexokinase catalyzes this phosphoryl group transfer. To start this reaction, ATP forms a complex with magnesium (II) ion and glucose binds to hexokinase. The magnesium-ATP complex then binds with the hexokinase-glucose complex and forms an intermediate (Zeng, et al. present a picture showing the interctions of brain hexokinase with ATP). are thought to be involved in binding the magnesium ion in the magnesium-ATP complex [4]. The hydroxyl group on the terminal phosphoryl group of the ATP molecule nucleophilically attacks carbon 6 on glucose. This produces glucose-6-phosphate still bound to hexokinase and ADP still in complex with magnesium ion [5]. Glucose-6-phosphate and the magnesium-ADP complex leave hexokinase. Glucose-6-phosphate and ADP are the products of this reaction. Hexokinase undergoes an induced-fit conformational change when it binds to glucose, which ultimately prevents the hydrolysis of ATP. It also experiences potent allosteric inhibition under physiological concentrations by its immediate products, glucose-6-phosphate [4]. This is a mechanism by which the influx of substrate into the glycolytic pathway is controlled.


Kinetics and Inhibition of HexokinaseKinetics and Inhibition of Hexokinase

Hexokinase activates glycoloysis by phosphorylating glucose. Since the phosphorylation of glucose to glucose-6-phosphate is the rate limiting step of glucose metabolism, hexokinase has a very important role in regulating healthy glucose levels in the human body [7]. Hexokinase has high affinity, thus a low Km, for glucose. Tissues where hexokinase is present use glucose at low blood serum levels.

G6P inhibits hexokinase by binding to the N-terminal domain(this is simple feedback inhibition). It competitively inhibits the binding of ATP [8]. If the cell is not using the G6P that it is making, then it stops making it. In this way, hexokinase can also slow down glycolysis. Hexokinase I is thought to be the "pacemaker" of glycolysis in brain tissue and red blood cells [4]. Inorganic phosphate allosterically relieves hexokinase of inhibition by G6P [8].










[edit] Additional Resources For additional information, see: Carbohydrate Metabolism


[edit] References 1.↑ Pollard-Knight D, Cornish-Bowden A. Mechanism of liver glucokinase. Mol Cell Biochem. 1982 Apr 30;44(2):71-80. PMID:7048063

2.↑ 2.0 2.1 Kamata K, Mitsuya M, Nishimura T, Eiki J, Nagata Y. Structural basis for allosteric regulation of the monomeric allosteric enzyme human glucokinase. Structure. 2004 Mar;12(3):429-38. PMID:15016359 doi:10.1016/j.str.2004.02.005

3.↑ Postic C, Shiota M, Magnuson MA. Cell-specific roles of glucokinase in glucose homeostasis. Recent Prog Horm Res. 2001;56:195-217. PMID:11237213

4.↑ Zeng C, Aleshin A, Hardie J, Harrison R, Fromm H. ATP-Binding site of Human Brain Hexokinase as Studied by Molecular Modeling and Site-Directed Mutagenesis. Biochem. 1996 Aug 6;35:13157-13164.

5.↑ hammes G, and Kochavi D. Studies of the Enzyme Hexokinase: Kinetic Inhibition by Products. Massachusetts Institute of Technology. 1961 Oct 5.

6.↑ Ralph E, Thomson J, Almaden J, Sun S. Glucose Modulation fo Glucokinase Activation by Small Molecules. Biochem. 2008 Feb 15;47:5028-5036.

7.↑ Pal P, and Miller B. Activating Mutations in the Human Glucokinase Gene Revealed by Genetic Selection. Biochem. 2008 Dec 3;48:814-816.

8.↑ Aleshin A, Malfois M, Liu X, Kim C, Fromm H, Honzatko R, Koch M, Svergun D. Nonaggregating Mutant of Recombinant Human Hexokinase I Exhibits Wild-Type Kinetics and Rod-like Conformations in Solution. Biochem. 1999 Apr 29;38:8359-8366.

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