scholarly journals Characterization of the Acetate Binding Pocket in the Methanosarcina thermophila Acetate Kinase

2005 ◽  
Vol 187 (14) ◽  
pp. 5059-5059 ◽  
Author(s):  
Cheryl Ingram-Smith ◽  
Andrea Gorrell ◽  
Sarah H. Lawrence ◽  
Prabha Iyer ◽  
Kerry Smith ◽  
...  
Keyword(s):  
Binding Pocket ◽  
Acetate Kinase ◽  
Methanosarcina Thermophila

scholarly journals Characterization of the Acetate Binding Pocket in the Methanosarcina thermophila Acetate Kinase

2005 ◽  
Vol 187 (7) ◽  
pp. 2386-2394 ◽  
Author(s):  
Cheryl Ingram-Smith ◽  
Andrea Gorrell ◽  
Sarah H. Lawrence ◽  
Prabha Iyer ◽  
Kerry Smith ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Binding Site ◽  
Binding Pocket ◽  
Acetate Kinase ◽  
Phosphoryl Group ◽  
Acetyl Phosphate ◽  
Hydrophobic Pocket ◽  
Methanosarcina Thermophila ◽  
Methyl Group ◽  
Substrate Binding Sites

ABSTRACT Acetate kinase catalyzes the reversible magnesium-dependent synthesis of acetyl phosphate by transfer of the ATP γ-phosphoryl group to acetate. Inspection of the crystal structure of the Methanosarcina thermophila enzyme containing only ADP revealed a solvent-accessible hydrophobic pocket formed by residues Val93, Leu122, Phe179, and Pro232 in the active site cleft, which identified a potential acetate binding site. The hypothesis that this was a binding site was further supported by alignment of all acetate kinase sequences available from databases, which showed strict conservation of all four residues, and the recent crystal structure of the M. thermophila enzyme with acetate bound in this pocket. Replacement of each residue in the pocket produced variants with Km values for acetate that were 7- to 26-fold greater than that of the wild type, and perturbations of this binding pocket also altered the specificity for longer-chain carboxylic acids and acetyl phosphate. The kinetic analyses of variants combined with structural modeling indicated that the pocket has roles in binding the methyl group of acetate, influencing substrate specificity, and orienting the carboxyl group. The kinetic analyses also indicated that binding of acetyl phosphate is more dependent on interactions of the phosphate group with an unidentified residue than on interactions between the methyl group and the hydrophobic pocket. The analyses also indicated that Phe179 is essential for catalysis, possibly for domain closure. Alignments of acetate kinase, propionate kinase, and butyrate kinase sequences obtained from databases suggested that these enzymes have similar catalytic mechanisms and carboxylic acid substrate binding sites.

Characterization of the Acyl Substrate Binding Pocket of Acetyl-CoA Synthetase†

Biochemistry ◽  
2006 ◽  
Vol 45 (38) ◽  
pp. 11482-11490 ◽  
Author(s):  
Cheryl Ingram-Smith ◽  
Barrett I. Woods ◽  
Kerry S. Smith
Keyword(s):  
Substrate Binding ◽  
Binding Pocket ◽  
Substrate Binding Pocket ◽  
Acetyl Coa

scholarly journals Structural and Functional Characterization of K339T Substitution Identified in the PB2 Subunit Cap-binding Pocket of Influenza A Virus

2013 ◽  
Vol 288 (16) ◽  
pp. 11013-11023 ◽  
Author(s):  
Yong Liu ◽  
Kun Qin ◽  
Geng Meng ◽  
Jinfang Zhang ◽  
Jianfang Zhou ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Functional Characterization ◽  
Binding Pocket

scholarly journals Characterization of the anti-apoptotic mechanism of Bcl-B

2003 ◽  
Vol 376 (1) ◽  
pp. 229-236 ◽  
Author(s):  
Dayong ZHAI ◽  
Ning KE ◽  
Haichao ZHANG ◽  
Uri LADROR ◽  
Mary JOSEPH ◽  
...  
Keyword(s):  
Strong Correlation ◽  
Transmembrane Domain ◽  
Binding Pocket ◽  
Over Expression ◽  
Bh3 Domain ◽  
Family Protein ◽  
Critical Residues ◽  
Differential Binding ◽  
Apoptotic Mechanism

Bcl-B protein is an anti-apoptotic member of the Bcl-2 family protein that contains all the four BH (Bcl-2 homology) domains (BH1, BH2, BH3 and BH4) and a predicted C-terminal transmembrane domain. Our previous results showed that Bcl-B binds Bax and suppresses apoptosis induced by over-expression of Bax; however, Bcl-B does not bind or suppress Bak. To explore the molecular basis for the differential binding and suppression of Bax and Bak, we studied the BH3 dimerization domains of Bax and Bak. Chimeric mutants of Bax and Bak were generated that swapped the BH3 domains of these pro-apoptotic proteins. Bcl-B associated with and blocked apoptosis induced by mutant Bak containing the BH3 domain of Bax, but not mutant Bax containing the BH3 domain of Bak. In contrast, Bcl-XL protein bound and suppressed apoptosis induction by Bax, Bak and both BH3-domain chimeras. A strong correlation between binding and apoptosis suppression was also obtained using a series of alanine substitutions spanning the length of the Bax BH3 domain to identify critical residues for Bcl-B binding. Conversely, using structure-based modelling to design mutations in the BH3-binding pocket of Bcl-B, we produced two Bcl-B mutants (Leu86→Ala and Arg96→Gln) that failed to bind Bax and that also were unable to suppress apoptosis induced by Bax over-expression. In contrast, other Bcl-B mutants that still bound Bax retained protective activity against Bax-induced cell death, thus serving as a control. We conclude that, in contrast with some other anti-apoptotic Bcl-2-family proteins, a strong correlation exists for Bcl-B between binding to pro-apoptotic multidomain Bcl-2 family proteins and functional apoptosis suppression.

scholarly journals Discovery and Characterization of a Distinct Cyclic Nucleotide Binding Pocket in HCN Channels

2014 ◽  
Vol 106 (2) ◽  
pp. 627a
Author(s):  
Anna Moroni ◽  
Marco Lolicato ◽  
Annalisa Bucchi ◽  
Cristina Arrigoni ◽  
Stefano Zucca ◽  
...  
Keyword(s):  
Cyclic Nucleotide ◽  
Nucleotide Binding ◽  
Binding Pocket ◽  
Hcn Channels ◽  
Cyclic Nucleotide Binding

scholarly journals Characterization of the ATPase activity of topoisomerase II from Leishmania donovani and identification of residues conferring resistance to etoposide

2005 ◽  
Vol 390 (2) ◽  
pp. 419-426 ◽  
Author(s):  
Tanushri Sengupta ◽  
Mandira Mukherjee ◽  
Aditi Das ◽  
Chhabinath Mandal ◽  
Rakhee Das ◽  
...  
Keyword(s):  
Atpase Activity ◽  
Topoisomerase Ii ◽  
Leishmania Donovani ◽  
Binding Pocket ◽  
Dna Topoisomerase ◽  
Terminal Domain ◽  
Specific Selectivity ◽  
Eukaryotic Dna ◽  
First Time

We have cloned and expressed the 43 kDa N-terminal domain of Leishmania donovani topoisomerase II. This protein has an intrinsic ATPase activity and obeys Michaelis–Menten kinetics. Cross-linking studies indicate that the N-terminal domain exists as a dimer both in the presence and absence of nucleotides. Etoposide, an effective antitumour drug, traps eukaryotic DNA topoisomerase II in a covalent complex with DNA. In the present study, we report for the first time that etoposide inhibits the ATPase activity of the recombinant N-terminal domain of L. donovani topoisomerase II. We have modelled the structure of this 43 kDa protein and performed molecular docking analysis with the drug. Mutagenesis of critical amino acids in the vicinity of the ligand-binding pocket reveals less efficient inhibition of the ATPase activity of the enzyme by etoposide. Taken together, these results provide an insight for the development of newer therapeutic agents with specific selectivity.

Sign in / Sign up

Export Citation Format

Share Document