GLUTAMINE SYNTHETASE

The current methods of X-ray diffraction have determined the structure of glutamine synthetase (GS) to be composed of a total of 43,692 non-hydrogen atoms within twelve subunits, which are arranged in two layers of six. The twelve identical subunits are arranged as the carbon atoms in two face-to-face benzene rings. Upon dissociation, these monomers become inactive because the active sites of bacterial GS are formed from pairs of adjacent subunits. GS as depicted above represents only one subunit of the entire enzyme which is complexed with glutamate within its active site. The active site is significant due to the main functional role of glutamine sythetase: it regulates the rate of glutamine synthesis in response to feedback inhibition and covalent inhibition.

Glutamine sythetase catalyzes the ATP-dependent amidation of glutamate to form glutamine. This specific reaction proceeds via a gamma-glutamyl-phosphate intermediate, and GS activity depends on the presence of divalent cations such as Mn²⁺. Glutamine is a major nitrogen donor in the biosynthesis of many organic nitrogen compounds such as purines, pyrimidines, and other amino acids.¹

Different Renderings of the Enzyme:

Colored Backbone Chain

Space Filling View

The icon below depicts the secondary structure of GS. In the image, the alpha helices are identified in pink, whereas the beta sheets are identified in gold.

Colored Cartoon View

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Strutural Characteristics:

In order to view the respective regions of positive and negative charge within this enzyme subunit, one must click on the icon below. In the image, the acidic residues are identified in magenta and the basic residues are identified in yellow.

Acidic/Basic Structure

In order to view the different degrees of hydrophobicity, one must click on the icon below. In the image, the polar amino acids are identified in red and the hydrophobic amino acids are identified in green blue.

Hydrophobicity

Solvent Accessible Surface Areas:

To view the solvent accessible surface areas of the enzyme, pay close attention to the directions below:

Right click on the image---->Options---->Dot surface---->Connolly/Richards Solvent (1.2A)

Specific Contact Sites--Hydrogen Bonding:

The image below as derived from Quanta/Charmm portrays the inherent hydrogen bonding within the active site of GS. The glutamate ligand is rendered as the ball and stick molecule. The two small grey spheres within this image represent the Mn²⁺ ions. As seen below, hydrogen bonding contributes to the ligand-enzyme complexion. The top carboxylic acid end of the ligand is hydrogen bonding to an arginine residue, and the amide group of the ligand is hydrogen bonding to glutamic acid residues of GS. The assumption concerning the bottom carboxylic acid end of glutamate is that it is hydrogen bonding to either arginine or histidine residues due to their presence within the active site.

The Active Site of Glutamine Synthetase:

Active Site Residues	Active Site Residues
Glu 131	His 269
Arg 321	Gly 265
Arg 339	Arg 359
Asp 50	Glu 471

Residues Surrounding Mn2+ Ions

Glu 357

His 269

Glu 129

Glu 131

Glu 220

Glu 212

The active site residues as listed in the table above are charged with the exception of glycine-265. The divalent cations (Mn²⁺) are surrounded by negatively charged glutamic acid residues with the exception of histidine-269. As discussed in the introduction, the divalent cations play an essential role in the catalyis of the substrate. GS couples the formation of glutamine with the hydrolysis of ATP into ADP and P_i. Research has also investigated the allosteric regulation of GS activity via feeback inhibition. Although there are a total of nine distinct feedback inhibitors, glycine, alanine and serine, as key indicators of amino acid metabolism within cells, are important regulators of GS activity because they compete with glutamate for binding at the active site.¹ The active site is essential for glutamine formation and feedback inhibition; however, it has no effect on such factors as covalent modification, which causes GS to be interconverted between the active and inactive forms.  

A Stimulating Mixed Rendering Image:

Ramachandran Plot:

The specific plot below depicts the phi and psi angles for glutamine synthetase. The colored regions indicate particularly favorable values for these angles. These favorable values correlate with the enzyme's inherent secondary structure as shown in the renderings above. More specifically, the dots within the purple region correspond to angles that can be attributed to alpha helices, whereas the dots within the yellow region correspond to angles that can be attributed to beta sheets.  

Minimized Structure of Glutamine Synthetase Using Quanta Charmm:

Breakdown of Energy Values: 

 The structural characteristics as depicted and described for glutamine synthetase provide for the unique function of catalyzing the formation of glutamine. As presented, GS is regulated by such factors as feedback inhibition and covalent modifications. This enzyme's active site contains metal ions and is capable of hydrogen bonding with the glutamate ligand, both of which play an essential role in catalytic reactions. The catalysis of glutamate to glutamine is coupled with the hydrolysis of ATP. The ramaschandran plot and the breakdown of energy values have also been presented in order to obtain a better understanding of this biologically interesting enzyme.