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1vm7

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Table of contents
  1. 1. Protein Summary
  2. 2. Ligand Summary
  3. 3. References

Title Crystal structure of Ribokinase (TM0960) from Thermotoga maritima at 2.15 A resolution. To be published
Site JCSG
PDB Id 1vm7 Target Id 282829
Molecular Characteristics
Source Thermotoga maritima msb8
Alias Ids TPS1249,TM0960, 3.40.1190.20, 84675 Molecular Weight 32652.69 Da.
Residues 299 Isoelectric Point 4.96
Sequence mflvisvvgssnidivlkvdhftkpgetqkaiemnvfpggkganqavtvakigekgcrfvtcignddys dllienyeklgitgyirvslptgrafievdktgqnriiifpganaelkkelidwntlsesdilllqnei pfettlecakrfngivifdpapaqgineeifqyldyltpnekeiealskdffgefltvekaaekflelg vknvivklgdkgvllvnknekkhfptfkvkavdttaagdvfngafavalsegknpeeavifgtaaaais vtrlgaqssipareeveaflknl
  BLAST   FFAS

Structure Determination
Method XRAY Chains 2
Resolution (Å) 2.15 Rfree 0.23539
Matthews' coefficent 2.34 Rfactor 0.18954
Waters 279 Solvent Content 47.13

Pathway

Reactions found in Metabolic Reconstruction for TM0960

Name: ribokinase reversible
Metabolic Subsystem: Ribose Metabolism
Reaction: : atp + rib-D <==> adp + h + r5p
Classification: EC:2.7.1.15
 
Name: Deoxyribokinase reversible
Metabolic Subsystem: Ribose Metabolism
Reaction: : atp + drib <==> 2dr5p + adp + h
Classification: EC:2.7.1.15
 

Ligand Information
Ligands
Metals

Jmol

 
Google Scholar output for 1vm7
1. An iterative knowledge_based scoring function for proteinprotein recognition
SY Huang, X Zou - Proteins: Structure, Function, and , 2008 - Wiley Online Library
 
2. Overexpression, purification and crystallographic analysis of a unique adenosine kinase from Mycobacterium tuberculosis
Y Wang, MC Long, S Ranganathan - Section F: Structural , 2005 - scripts.iucr.org
 
3. Structure of Methanocaldococcus jannaschii nucleoside kinase: an archaeal member of the ribokinase family
L Arnfors, T Hansen, P Schonheit - Section D: Biological , 2006 - scripts.iucr.org
 

Protein Summary

The gene TM0960 from Thermotoga maritima encodes ribokinase from pfkB-like superfamily of carbohydrate kinases PF00294 COG0524.  Ribokinase catalyses the phosphorylation of ribose to ribose-5-phosphate using ATP. This reaction is the first step in the metabolism of ribose.  Ribokinase traps ribose within the cell after uptake and also prepares the sugar for use in the synthesis of nucleotides and histidine, and for entry into the pentose phosphate pathway. This family also includes a variety of carbohydrate and pyrimidine kinases.  This enzyme is a part of the thiamine pyrophosphate (TPP) synthesis pathway. TPP is an essential cofactor for many enzymes [Ref] .  Ribokinase is dimeric in solution.

 

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TM0960: Experimentally confirmed a ribokinase

The Thermotoga maritima TM0960 ORF (1VM7) was annotated as a putative ribokinase based on sequence and structural comparisons. TM0960 is similar in sequence to ribokinase from E. coli (1RK2), human (2FV7) and Mycobacterium tuberculosis (3GO6). Ribokinase has a tertiary structure consisting of large αβα sandwich domain and a smaller lid domain via a conserved “GG switch” motif.  The two glycine residues comprising the switch undergo torsional rotation upon substrate binding, which “closes” the lid domain over the substrate-binding site, also present in the large sandwich domain. This lid closure initiates a series of conformational changes, ultimately resulting in formation of the enzyme’s active site and catalysis of the family-wide conserved reaction1.

Michaelis-Menten kinetics data support the function of TM0960 as a ribokinase. TM0960 catalyzes the conversion of D-ribose (KM=0.92 mM,kcat =1.1 sec-1) and ATP (KM=0.018 μM,kcat=0.26 sec-1) to D-ribose-5-phosphate and ADP. TM0960 displays less activity in the presence of D-ribose compared to structural homologues E.coli ribokinase (1RK2) (KM=0.18 mM)2 and human ribokinase (2FV7) (KM=0.279 mM, kcat=86.4 sec-1)3. Experimental temperature (22°C) may have modulated TM0960 activity, as the ideal temperature for the hyperthermophilic Thermatoga maritima is 80°C. Substrate specificity was investigated by testing enzymatic activity with the D- ribose, D -glucose, D -fructose, and 2-deoxy-D -ribose as phosphate acceptors and ATP, GTP, and UTP as phosphate donors. TM0960 is highly specific for D -ribose as only D -ribose was able to be phosphorylated; conversely, ATP, GTP, and UTP served equally well as phosphate donors (Figure 2). Substrate inhibition was observed when ribose was above 2 mM, similar to the inhibition of human ribokinase by D-ribose and ATP at concentrations about 0.5 mM and 0.3 mM, respectively4. TM0960 requires divalent cations for activity, though the requirement is non-specific; Mg2+, Mn2+, and Co2+, all produced comparable specific activities while the presence of Zn2+ resulted in roughly 50% of optimal activity. Despite its role as an essential activator of TM0960, Mg2+ was shown to inhibit the enzyme’s activity when present at concentrations above 3 mM. Human and E. coli ribokinases also demonstrate ribokinase inhibition above 2 mM and 5 mM Mg2+, respectively3,4.

 

1Sigrell, J., Cameron, A., and Mowbray, S. (1999) Induced fit on sugar binding activates ribokinase. J. Molecular Biology, 290(5), 1009-1018.

2Maj, M. C., and Gupta, R. S. (2001) The effect of inorganic phosphate on the activity of bacterial ribokinase. J. Protein Chemistry, 20(2), 139-144.

3Chuvikovsky D. V., Esipov R. S., Skoblov Y. S., Chupova L. A., Muravyova T. I., Miroshnikov A. I., Lapinjoki S, and Mikhailopulo I.A. (2006). Ribokinase from E. coli: Expression, purification, and substrate specificity. Bioorganic & Medicinal Chemistry, 14(18), 6327-6332.

4Park J., van Koeverden P., Singh B., and Gupta R. S. (2007). Identification and characterization of human ribokinase and comparison of its properties with E-coli ribokinase and human adenosine kinase. FEBS Letters, 581(17), 3211-3216.

BioLED contributors: Kaitlin Bailey, Carol Price, Karlyn Pollack, Laura Ramsey, Jonathan Armstrong, Vietvuong Vo, Steve Farruggio, Tomasz Kabzinski, Cameron Mura, Linda Columbus. Funded by NSF DUE 1044858.

Ligand Summary



References

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References

 

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Files (2)

FileSizeDateAttached by 
TM0960 TOPSAN submission figure 1.png
TM0960 Figure 1 HW plot of ribokinase kinetics
34.88 kB15:58, 10 Jun 2013cawpriceActions
 TM0960 TOPSAN submission figure 2.png
TM0960 Figure 2 phosphate donor specficity
32.18 kB15:56, 10 Jun 2013cawpriceActions
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