Proteins from Mycobacterium tuberculosis: Difference between revisions
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<StructureSection load='' size='450' side='right' scene='80/801748/Cv/1' caption=''> | <StructureSection load='' size='450' side='right' scene='80/801748/Cv/1' caption=''> | ||
===Novel T9 loop interaction of Filamenting Temperature-sensitive mutant Z from ''Mycobacterium tuberculosis''<ref>doi 10.1107/S2053230X19004618</ref>=== | |||
As of 2017, tuberculosis has infected 1.7 billion people (23% of the world's population) and has caused 10 million deaths. ''Mycobacterium tuberculosis'' (Mtb) is quickly evolving, and new strains are classified as multi-drug resistant. Thus, the development and discovery of new drugs to combat Mtb is vital to combat the drug-resistant strains. Filamenting temperature-sensitive mutant Z (FtsZ), an important protein involved in cell-division is key for the survival of Mtb. Here, we have solved the crystal structure of MtbFtsZ that exhibit an inter-subunit that plays a biological role in the GTPase activity of MtbFtsZ and have elucidated a novel conformation, involving the T9 loop and the nucleotide binding pocket that breaks up the GTPase active site. This novel conformation can serve as basis for the development of the novel drugs to combat tuberculosis. | |||
<scene name='81/813404/Cv/2'>Asymmetric unit contains 6 protomers that form a dimer of trimers</scene> ([[5v68]]). Protomers A and C shown in blue, protomer B shown in gray, protomers D and F shown in red, protomer E shown in cyan, GDP are represented by orange spheres, and PO4 by green spheres. <scene name='81/813404/Cv/4'>Trimer ABC superimposed onto DEF</scene>. Same color scheme as in previous scene. | |||
Comparison of other MtbFtsZ structures with trimer ABC from [[5v68]] reveals that protomers A and B superimposed well but the conformation of protomer C is vastly different. <scene name='81/813404/Cv/23'>Structure 5V68 superimposed onto 4KWE</scene>. Nucleotide for both is GDP. Color scheme is as follows, [[5v68]] blue, [[4kwe]] yellow, nucleotides orange spheres, Glu231 green spheres, phosphate red spheres. <scene name='81/813404/Cv/24'>Click here to see animation of this scene</scene>. <jmol><jmolButton> | |||
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</jmolButton></jmol> The trimer ABC from [[5v68]] is to a curved MtbFtsZ protofilament (PDB [[4kwe]]). Based on an r.m.s.d. of 1.2Å between protomers AB of [[5v68]] and protomers CB of [[4kwe]], protomers A and B from [[5v68]] are similar with protomers C and B of [[4kwe]]. <scene name='81/813404/Cv/25'>Glu231 (shown as green sphere) from protomer C of [[5v68]] and Glu231 from protomer A of [[4kwe]] are far apart</scene>. | |||
<scene name='81/813404/Cv/26'>5V68 with bound GDP superimposed onto 2Q1Y with bound GTP</scene>. Color scheme is as follows, [[5v68]] blue, [[2q1y]] green, nucleotides orange spheres, Glu231 green spheres, phosphate red spheres. <scene name='81/813404/Cv/27'>Click here to see animation of this scene</scene>. <jmol><jmolButton> | |||
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</jmolButton></jmol> To generate this trimer ([[2q1y]]), A1 A2 A3, the crystal symmetry molecules neighboring chain A2 that exhibit the inter-subunit interface were used. Only chains A1 and A2 were used for the superimposition. Again, protomers A and B from present study are similar with the top two protomers A1 and A2 from [[2q1y]] with an r.m.s.d of 2.1Å. <scene name='81/813404/Cv/29'>The distance of their respective Glu231</scene> is approximately 63Å. | |||
<scene name='81/813404/Cv/21'>4KWE with bound GDP superimposed onto 2Q1Y with bound GTP</scene>. Color scheme is as follows, [[4kwe]] yellow, [[2q1y]] green, nucleotides orange spheres, Glu231 green spheres. <scene name='81/813404/Cv/22'>Click here to see animation of this scene</scene>. <jmol><jmolButton> | |||
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</jmolButton></jmol> Superimposition of [[4kwe]] and [[2q1y]] reveals an r.m.s.d. of 2.8Å between the trimers and a 12.5Å <scene name='81/813404/Cv/30'>distance between their respective Glu231’s of A and A3</scene>, showing that these structures are relatively similar. The distances between Glu231 of [[5v68]] from protomer C is far greater than 12.5Å and the interaction with the middle protomers of [[4kwe]] and [[2q1y]] with protomer C of [[5v68]] involve the T9 loop residue Glu231, which is not the case in the other structures. | |||
<scene name='81/813404/Cv1/3'>Crystal structure of protomers A and B of 5V68</scene> (gray). <scene name='81/813404/Cv1/6'>A zoom-in of the inter-subunit interface with bound GDP</scene> (orange). Helix H11 is shown in green. Helices η1, H7, and loop T6 are shown in blue. Switch I (T3 loop) is disordered surrounded by a cloud. Switch II (sH2) is shown in magenta. The top two protomers of all three structures (AB for [[5v68]], A1 A2 for [[2q1y]], and CB for [[4kwe]]) all exhibit a similar inter-subunit interface. This interaction between protomers A and B of 5V68 involve the T6 and T7 loops, helices: H11, η1, and H7. Protomer A of our structure “sits” on the helices η1, H7 and loop T6 (all shown in blue) from protomer B. This brings the T7 loop’s (shown in red) residues <scene name='81/813404/Cv1/9'>Asn205, Asp207 and Asp210</scene> of protomer A of [[5v68]] within 16Å of GDP from protomer B, forming the GTPase active site. The T3 loop is disordered or in its OFF position (surrounded by magenta cloud). Helix sH2 is also OFF because there is no hydrogen network (shown in magenta). | |||
<scene name='81/813404/Cv1/11'>Crystal structure of protomer B and C of 5V68</scene>. Protomer B is shown in gray, the T9 loop in brown, T11 in cyan; and the N-terminal of protomer C is shown in green, C-terminal blue, helix H8 yellow, the T7 loop red, and the switches in magenta. <scene name='81/813404/Cv1/12'>PO4 binding site</scene>. Glu231 from protomer B is shown in brown, while Glu274 from protomer is in gray. The residues in green are from protomer C. | |||
===Substrate analogue complex structure of ''Mycobacterium tuberculosis'' decaprenyl diphosphate synthase<ref>doi 10.1107/S2053230X19001213</ref> === | |||
Rv1086 produces Ω-''E,Z''-farnesyl diphosphate (''EZ''-FPP, C15) from geranyl diphosphate (GPP, C10) and isopentenyl diphosphate (IPP, C5) that is used by Rv2361c for further elongation to form decaprenyl diphosphate (DPP, C50). In this structure we have substrate analogues of GPP and IPP as well as the essential Mg ion bound to Rv2361 (''i.e.'' MtDPPS) in a productive mode. GPP binds to S1 site and IPP binds to S2 site. The hydrocarbon chains are joined head-to-tail to form a 5-carbon longer product. Meanwhile, we also have the GPP analogue bound in alternate conformations. The varying interactions of this substrate with Asp76 from one subunit and Arg292 from another may account for the transition pathway from S2 site to S1 site after each cycle of elongation reaction. So the enzyme can proceed to the next cycle of catalysis. | |||
<scene name='80/806392/Cv/8'>Overall structure of MtDPPS</scene>. The two monomers in an asymmetric unit of the MtDPPS crystal are shown as ribbon diagrams. The β-strands are named A-F and the α-helices numbered 1-7 from N to C terminus. They are colored yellow/red for one subunit and magenta/cyan for the other. | |||
The reaction catalyzed by Rv2361c (or ''M. tuberculosis'' DPP synthase, MtDPPS) is very similar to that of undecaprenyl diphosphate synthase (UPPS), except for the chain length of the final product (C<sub>50</sub> vs C<sub>55</sub>) and the starting allylic substrate (''EZ''-FPP vs ''EE''-FPP). In fact, most ''cis''-PTs share a common dimeric architecture, and the conserved S1 and S2 sites for substrate binding are located near the subunit interface. The starting allylic substrate is bound to the S1 site and the homoallylic substrate to be incorporated is bound to the S2 site. An invariant aspartate residue plays a central role in the catalysis by coordinating the Mg2+-bound substrates. The head-to-tail coupling reaction of ''cis''-PT proceeds through a concerted pathway similar to the ionization-condensation-elimination mechanism of ''trans''-PT. After the new C-C bond formation, the pyrophosphate leaves the S1 site along with Mg2+, and the resulting prenyl diphosphate switches from the S2 site to the S1 site (see static image below). | |||
[[Image:RxnPathway.png|left|400px|thumb| The reaction pathway of ''cis''-prenyltransferase in general. C<sub>5n</sub> stands for a hydrocarbon group of n prenyl units.]] | |||
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<scene name='80/806392/Cv1/6'>Binding modes of substrate analogues</scene>. The MtDPPS dimer is superimposed on itself with the two polypeptide chains switched. The protein is colored cyan/green in one dimer and pink/yellow in the other, and so are the side chains and the ligands, which are shown as stick models. Mg and water molecules are shown as spheres, and the coordinate bonds as dashed lines. Location of the S1 and S2 site as well as the nearby helices α1/α2 and strand βB are also indicated. When the S1 and S2 substrates and Mg are properly bound for catalysis, the <scene name='80/806392/Cv1/9'>Asp76 side chain not only binds directly to Mg but also to a coordinating water molecule</scene>. The <scene name='80/806392/Cv1/10'>same water is hydrogen bonded to the side chain of Arg292*, which also binds to the other Mg-bound water</scene> (the asterisks denote residues from the counter-subunit in a dimer). <scene name='80/806392/Cv1/11'>In the absence of both the S2 substrate and Mg, Arg292* turns to bind directly to Asp76</scene>, which is no longer engaged in Mg-coordination. The <scene name='80/806392/Cv1/12'>side chain of Arg292* binds to the β-phosphate of the S1 substrate in this conformation</scene>, and in the other it is also close to the β-phosphate of the S2 substrate. After the formation of new ''cis''-double bond, the S1 pyrophosphate dissociates as an Mg complex, and Arg292* binds to the β-phosphate of the product and transfers it to the S1 site (see static image below). While the five-carbon longer hydrocarbon tail needs structural rearrangements to fit into the S1 pocket, the diphosphate moiety may be disposed like those of the GSPP conformers before it assumes a productive binding mode for the next cycle of reaction. | |||
[[Image:Transloc.png|left|400px|thumb|In this schematic diagram, the side chains of Asp76 and Arg292 are colored dark red and dark blue. The three subsites for the alternative binding modes of the S1 substrate are indicated by green curves. Other bonds, including the Mg-coordinates, are in black. R<sub>n</sub> stands for a group of n consecutive isoprene units (C<sub>5n</sub>).]] | |||
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=== The structure of ''Mycobacterium tuberculosis'' HtrA reveals an auto-regulatory mechanism<ref>doi 10.1107/S2053230X18016217</ref> === | === The structure of ''Mycobacterium tuberculosis'' HtrA reveals an auto-regulatory mechanism<ref>doi 10.1107/S2053230X18016217</ref> === | ||
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<scene name='80/801748/Cv/25'>A comparison of the active site conformation in different HtrA enzymes with bovine trypsin</scene> (<span style="color:yellow;background-color:black;font-weight:bold;">''M. tuberculosis'' HtrA active site residues are in yellow</span>). The active site residues from bovine trypsin and proteases belonging to the HtrA family from different species were superposed with ''M. tuberculosis'' ΔTM HtrA. Two well characterized HtrA proteases (''M. tuberculosis'' PepD (PDB ID: [[2z9i]]; <span style="color:salmon;background-color:black;font-weight:bold;">colored in salmon</span>), E. coli DegS (PDB ID: [[1soz]]; <span style="color:cyan;background-color:black;font-weight:bold;">colored in cyan</span>)) provided a basis for this comparison alongside bovine trypsin structures. Among these, one is a complex with phenylmethylsulfonyl fluoride (PMSF) (PDB ID: [[1pqa]]; <span style="color:dodgerblue;background-color:black;font-weight:bold;">colored in dodgerblue</span>) providing a reference for a covalently linked ligand to active site Ser-OH. The other representative model for a substrate bound form is the trypsin-peptide complex (AAPK) (PDB ID: [[2agg]]; <span style="color:violet;background-color:black;font-weight:bold;">colored in violet</span>). This structure provides a representation of the oxyanion hole wherein the peptide is bound to the active site Ser-OH providing a structural snapshot of the acyl enzyme intermediate. In both examples, the active site Histidine is flipped with χ<sup>1</sup> of 80.7° and -166.8° in the case of peptide bound ([[1pqa]]) or 89.4° and -174.8° in the case of the PMSF complex ([[2agg]]).<ref name="Ash">PMID:10984533</ref><ref name="Radisky">PMID:16636277</ref> For comparison, the Histidine rotamers with χ<sup>1</sup> of 80.7° and 89.4° represent the canonical catalytic triad alongside the active site Asp and Ser. Of note, that the χ<sup>1</sup> of His270 of ''M. tuberculosis'' HtrA is -80.9 leading a distorted catalytic triad. This conformation of the catalytic triad in ''M. tuberculosis'' HtrA thus represents either an inactive state or a distorted conformation mimicking Histidine flipping in the acylation step of catalysis. | <scene name='80/801748/Cv/25'>A comparison of the active site conformation in different HtrA enzymes with bovine trypsin</scene> (<span style="color:yellow;background-color:black;font-weight:bold;">''M. tuberculosis'' HtrA active site residues are in yellow</span>). The active site residues from bovine trypsin and proteases belonging to the HtrA family from different species were superposed with ''M. tuberculosis'' ΔTM HtrA. Two well characterized HtrA proteases (''M. tuberculosis'' PepD (PDB ID: [[2z9i]]; <span style="color:salmon;background-color:black;font-weight:bold;">colored in salmon</span>), E. coli DegS (PDB ID: [[1soz]]; <span style="color:cyan;background-color:black;font-weight:bold;">colored in cyan</span>)) provided a basis for this comparison alongside bovine trypsin structures. Among these, one is a complex with phenylmethylsulfonyl fluoride (PMSF) (PDB ID: [[1pqa]]; <span style="color:dodgerblue;background-color:black;font-weight:bold;">colored in dodgerblue</span>) providing a reference for a covalently linked ligand to active site Ser-OH. The other representative model for a substrate bound form is the trypsin-peptide complex (AAPK) (PDB ID: [[2agg]]; <span style="color:violet;background-color:black;font-weight:bold;">colored in violet</span>). This structure provides a representation of the oxyanion hole wherein the peptide is bound to the active site Ser-OH providing a structural snapshot of the acyl enzyme intermediate. In both examples, the active site Histidine is flipped with χ<sup>1</sup> of 80.7° and -166.8° in the case of peptide bound ([[1pqa]]) or 89.4° and -174.8° in the case of the PMSF complex ([[2agg]]).<ref name="Ash">PMID:10984533</ref><ref name="Radisky">PMID:16636277</ref> For comparison, the Histidine rotamers with χ<sup>1</sup> of 80.7° and 89.4° represent the canonical catalytic triad alongside the active site Asp and Ser. Of note, that the χ<sup>1</sup> of His270 of ''M. tuberculosis'' HtrA is -80.9 leading a distorted catalytic triad. This conformation of the catalytic triad in ''M. tuberculosis'' HtrA thus represents either an inactive state or a distorted conformation mimicking Histidine flipping in the acylation step of catalysis. | ||
=== The crystal structure of Acyl carrier protein synthase (AcpS) from ''Mycobacterium tuberculosis'' (''Mtb'') === | === The crystal structure of Acyl carrier protein synthase (AcpS) from ''Mycobacterium tuberculosis'' (''Mtb'')<ref>PMID:19733180</ref> === | ||
The crystal structure of AcpS from (''Mtb'') was solved at 1.95 Å ([[3hqj]]). It crystallized as one <scene name='3hqj/Trimer/2'>monomer</scene> per asymmetric unit. Since ''Mtb'' AcpS has biologically active trimeric arrangement, <scene name='3hqj/Trimer/3'>AcpS trimer</scene> (in <span style="color:lime;background-color:black;font-weight:bold;">green</span>, <font color='blue'><b>blue</b></font>, and <scene name='3hqj/Trimer/3'>AcpS trimer</scene> (in <span style="color:orange;background-color:black;font-weight:bold;">orange</span>) was constructed using the 3-fold crystallographic symmetry in the ''P''23 space group. | The crystal structure of AcpS from (''Mtb'') was solved at 1.95 Å ([[3hqj]]). It crystallized as one <scene name='3hqj/Trimer/2'>monomer</scene> per asymmetric unit. Since ''Mtb'' AcpS has biologically active trimeric arrangement, <scene name='3hqj/Trimer/3'>AcpS trimer</scene> (in <span style="color:lime;background-color:black;font-weight:bold;">green</span>, <font color='blue'><b>blue</b></font>, and <scene name='3hqj/Trimer/3'>AcpS trimer</scene> (in <span style="color:orange;background-color:black;font-weight:bold;">orange</span>) was constructed using the 3-fold crystallographic symmetry in the ''P''23 space group. | ||
The 3′,5′-ADP moieties of the [http://en.wikipedia.org/wiki/Coenzyme_A coenzyme A] (<font color='magenta'><b>CoA, colored magenta</b></font>), are positioned in the cleft between each of two monomers forming three active sites within AcpS trimer. The <scene name='3hqj/Trimer/5'>active site</scene> is formed by the residues <span style="color:lime;background-color:black;font-weight:bold;">D9 (highly conserved), E58, L62, and S65</span> from monomer <span style="color:lime;background-color:black;font-weight:bold;">A</span> and by <span style="color:orange;background-color:black;font-weight:bold;">R92, P93, R53, H116, and T115</span> from the neighboring monomer <span style="color:orange;background-color:black;font-weight:bold;">B</span>. The residues labeled and shown as sticks (A and B in the brackets point on the name of the monomer). Hydrogen bonds are shown as dashed lines with interatomic distances in Å. The magnesium (Mg) atoms are shown in spacefill representation and colored in <span style="color:cyan;background-color:black;font-weight:bold;">cyan</span>. The <font color='magenta'><b>CoA</b></font> is shown in stick representation and colored <font color='magenta'><b>magenta</b></font>. <font color='blue'><b>Nitrogen</b></font> and <font color='red'><b>oxygen</b></font> atoms of the CoA 3′,5′-ADP moiety and of the active site resudues are colored <font color='blue'><b>blue</b></font> and <font color='red'><b>red</b></font>, respectively. | The 3′,5′-ADP moieties of the [http://en.wikipedia.org/wiki/Coenzyme_A coenzyme A] (<font color='magenta'><b>CoA, colored magenta</b></font>), are positioned in the cleft between each of two monomers forming three active sites within AcpS trimer. The <scene name='3hqj/Trimer/5'>active site</scene> is formed by the residues <span style="color:lime;background-color:black;font-weight:bold;">D9 (highly conserved), E58, L62, and S65</span> from monomer <span style="color:lime;background-color:black;font-weight:bold;">A</span> and by <span style="color:orange;background-color:black;font-weight:bold;">R92, P93, R53, H116, and T115</span> from the neighboring monomer <span style="color:orange;background-color:black;font-weight:bold;">B</span>. The residues labeled and shown as sticks (A and B in the brackets point on the name of the monomer). Hydrogen bonds are shown as dashed lines with interatomic distances in Å. The magnesium (Mg) atoms are shown in spacefill representation and colored in <span style="color:cyan;background-color:black;font-weight:bold;">cyan</span>. The <font color='magenta'><b>CoA</b></font> is shown in stick representation and colored <font color='magenta'><b>magenta</b></font>. <font color='blue'><b>Nitrogen</b></font> and <font color='red'><b>oxygen</b></font> atoms of the CoA 3′,5′-ADP moiety and of the active site resudues are colored <font color='blue'><b>blue</b></font> and <font color='red'><b>red</b></font>, respectively. | ||
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The ''B. subtilis'' AcpS trimer ([[1f80]]) <scene name='3hqj/Acp/2'>binds</scene> three molecules of the acyl carrier protein (ASP). The interactions between ''B. subtilis'' AcpS and ACP are predominantly <scene name='3hqj/Acp/3'>electrostatic</scene>. The ''B. subtilis'' AcpS (white) is shown in spacefill representation, the agrinines, lysines, and histidines are colored <font color='blue'><b>blue</b></font>, while aspartates and glutamates are colored <font color='red'><b>red</b></font>. The ACP molecule (<span style="color:lime;background-color:black;font-weight:bold;">green</span>) is shown in ribbon representation with aspartates and glutamates as sticks and colored <font color='red'><b>red</b></font>. The ''B. subtilis'' AcpS has large <scene name='3hqj/Acp/4'>electropositive interface</scene> with ASP. <scene name='3hqj/Acp/5'>Electrostatic representation</scene> of ''Mtb'' AcpS surface using the similar orientation as ''B. subtilis'' AcpS, shows a moderate electronegative nature in the putative ACP binding site near the <font color='red'><b>ASP 15</b></font>. The ''Mtb'' ASPM structure ([[1klp]], corresponding to ACP) demonstrates considerably lower negative charge. So, the electrostatic interactions between ''Mtb'' AcpS and ASPM are, probably, less important. | The ''B. subtilis'' AcpS trimer ([[1f80]]) <scene name='3hqj/Acp/2'>binds</scene> three molecules of the acyl carrier protein (ASP). The interactions between ''B. subtilis'' AcpS and ACP are predominantly <scene name='3hqj/Acp/3'>electrostatic</scene>. The ''B. subtilis'' AcpS (white) is shown in spacefill representation, the agrinines, lysines, and histidines are colored <font color='blue'><b>blue</b></font>, while aspartates and glutamates are colored <font color='red'><b>red</b></font>. The ACP molecule (<span style="color:lime;background-color:black;font-weight:bold;">green</span>) is shown in ribbon representation with aspartates and glutamates as sticks and colored <font color='red'><b>red</b></font>. The ''B. subtilis'' AcpS has large <scene name='3hqj/Acp/4'>electropositive interface</scene> with ASP. <scene name='3hqj/Acp/5'>Electrostatic representation</scene> of ''Mtb'' AcpS surface using the similar orientation as ''B. subtilis'' AcpS, shows a moderate electronegative nature in the putative ACP binding site near the <font color='red'><b>ASP 15</b></font>. The ''Mtb'' ASPM structure ([[1klp]], corresponding to ACP) demonstrates considerably lower negative charge. So, the electrostatic interactions between ''Mtb'' AcpS and ASPM are, probably, less important. | ||
=== Drug resistance mechanism of PncA in ''Mycobacterium Tuberculosis'' <ref>doi 10.1080/07391102.2012.759885</ref>=== | |||
Tuberculosis continues to be a global health threat. Pyrazinamide (PZA) is an important first-line drug in multidrug-resistant tuberculosis treatment. The emergence of strains resistant to pyrazinamide represents an important public health problem, as both first- and second-line treatment regimens include pyrazinamide. It becomes toxic to ''Mycobacterium tuberculosis'' when converted to pyrazinoic acid by the <scene name='Journal:JBSD:11/Cv/5'>bacterial pyrazinamidase (PncA) enzyme</scene>. PZA resistance is caused mainly by the loss of enzyme activity by mutation, the mechanism of resistance is not completely understood. In our studies, we analysed three mutations (D8G, S104R and C138Y) of PncA which are resistance for <scene name='Journal:JBSD:11/Cv/6'>PZA</scene>. Binding pocket analysis solvent accessibility analysis, molecular docking and interaction analysis were performed to understand the interaction behaviour of mutant enzymes with PZA. Molecular dynamics simulations were conducted to understand the three dimensional conformational behaviour of <scene name='Journal:JBSD:11/Cv/3'>native</scene> and mutants PncA. Our analysis clearly indicates that the mutation (<scene name='Journal:JBSD:11/Cv/8'>D8G</scene>, <scene name='Journal:JBSD:11/Cv/9'>S104R</scene> and <scene name='Journal:JBSD:11/Cv/10'>C138Y</scene>) in PncA is responsible for rigid binding cavity which in turns abolishes conversion of PZA to its active form and is the sole reason for PZA resistance. Excessive hydrogen bonding between PZA binding cavity residues and their neighboring residues are the reason of rigid binding cavity during simulation. We present an exhaustive analysis of the binding-site flexibility and its 3D conformations that may serve as new starting points for structure-based drug design and helps there researchers to design new inhibitor with consideration of rigid criterion of binding residues due to mutation of this essential target. | |||
=== Enoyl-Acyl-Carrier Protein Reductase <ref>PMID:19130456</ref>=== | |||
[[Enoyl-Acyl-Carrier Protein Reductase]] is a target of anti-bacterial drugs such as triclosan (TCL). These drugs are used against tuberculosis infection. <scene name='43/434541/Cv/10'>Enoyl-Acyl-Carrier Protein Reductase is a tetramer</scene> (PDB code [[3fne]]). InhA ENR <scene name='43/434541/Cv/11'>active site contains NAD and triclosan derivative</scene>. | |||
=== Crystal structure of the essential biotin-dependent carboxylase AccA3 from Mycobacterium tuberculosis<ref>pmid 28469974</ref> === | |||
Biotin-dependent acetyl-CoA carboxylases catalyze the committed step in type II fatty acid biosynthesis, the main route for production of membrane phospholipids in bacteria, and are considered a key target for antibacterial drug discovery. Here we describe the first structure of AccA3, an essential component of the acetyl-CoA carboxylase system in ''Mycobacterium tuberculosis'' (MTb). The structure, sequence comparisons, and modeling of ligand-bound states reveal that the ATP cosubstrate-binding site shows distinct differences compared to other bacterial and eukaryotic biotin carboxylases, including all human homologs. This suggests the possibility to design MTb AccA3 subtype-specific inhibitors. | |||
''Mycobacterium tuberculosis'' <scene name='76/763765/Cv/2'>AccA3 adopts the ATPgrasp superfamily fold</scene>, and crystallized as a <scene name='76/763765/Cv/3'>dimer in the asymmetric unit</scene>. <scene name='76/763765/Cv/4'>The ordered structure of domain B is missing in chain B</scene>. | |||
Previous structures have shown defined ‘open’ and ‘closed’ states of the B-domain<ref>pmid 19213731</ref><ref>pmid 7915138</ref>. In addition, the biotin carboxylase domain of pyruvate carboxylase from ''Bacillus thermodenitrificans'' displays what appears to be an intermediate, but defined, conformation <ref>pmid 17642515</ref>. In the current structure, however, while <scene name='76/763765/Cv/6'>protomer A</scene> represents the previously observed ‘closed’ state, <scene name='76/763765/Cv/7'>protomer B</scene> represent a different structural state where no conformation is present in high enough occupancy to be possible to reliably model. MTb AccA3, subunit A (blue) and subunit B (yellow), unbound BDC from Escherichia coli (gray) (PDB [[1bnc]]). Based on the location of the segment of positive difference density relative to protomer B, it is, however, clear that the location of the B-domain in the partially occupied structural state that gives rise to this density is not the same as either the previously described ‘closed’ or ‘open’ states. Rather, the density suggests an even more extended conformation of the B-domain relative to the rest of the protein. Together, the most likely interpretation of the combined structural data of biotin-dependent carboxylases is that the B-domain is dynamic over a continuum of conformations, or several defined conformations. | |||
<scene name='76/763765/Cv/8'>Structural model</scene> of biotin and ADP binding in MTb AccA3 based on the biotin and ADP-bound ''Escherichia coli'' BDC (PDB [[3g8c]]). Substrate-bridging loop of ''MTb'' AccA3 rendered in pink and ''E. coli'' BDC in cyan. | |||
===[[Mycobacterium tuberculosis ArfA Rv0899]]=== | |||
</StructureSection> | </StructureSection> | ||