Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon -2

<p><b>Copyright information:</b></p><p>Taken from "Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon "</p><p></p><p>Molecular Microbiology 2007;65(3):700-713.</p><p>Published online 01 Aug 2007</p><p>PMCID:PMC1976387.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p

AIF2/5B stimulates the interaction of met-tRNAi with the archaeal 30S subunits

<p><b>Copyright information:</b></p><p>Taken from "Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon "</p><p></p><p>Molecular Microbiology 2007;65(3):700-713.</p><p>Published online 01 Aug 2007</p><p>PMCID:PMC1976387.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p> The amount of ribosome-bound met-tRNAi was determined by electrophoresis of the incubation mixtures on non-denaturing gels (see ). The autoradiography of the original gel is shown on the right. Lane 1: [S]Met-tRNAi without ribosomes; lane 2: [S]Met-tRNAi and 70S ribosomes without factors; lanes 3–5: [S]Met-tRNAi and 70S ribosomes with increasing amounts (10, 30, 50 pmol) of IF2; lanes 6–8: [S]Met-tRNAi and 70S ribosomes with increasing amounts (10, 30, 50 pmol) of aIF2/5B. The curve on the left is obtained by quantifying with a Molecular Dynamics Phosphoimager the radioactivity retained in the bands as shown in the panel on the right. Squares: increasing amounts of aIF2/5B; circles: Increasing amounts of bacterial IF2

Interaction of native and chimeric IF2 factors with archaeal and bacterial ribosomal subunits

<p><b>Copyright information:</b></p><p>Taken from "Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon "</p><p></p><p>Molecular Microbiology 2007;65(3):700-713.</p><p>Published online 01 Aug 2007</p><p>PMCID:PMC1976387.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p> The ribosome-bound fraction of the various proteins was determined as described in . Circle: recombinant aIF2/5B; diamond: recombinant IF2; triangle: BaSu1 chimera; square: BaSu2 chimera

Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon -6

<p><b>Copyright information:</b></p><p>Taken from "Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon "</p><p></p><p>Molecular Microbiology 2007;65(3):700-713.</p><p>Published online 01 Aug 2007</p><p>PMCID:PMC1976387.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p>synthesized in a translation systems programmed with increasing amounts of aIF2/5B mRNA and fixed amounts of a leaderless (α-fucosidase) or a leadered (104) mRNA. Lane 1: 10 pmol of α-fuc or 10 pmol of 104 mRNA in the absence of aIF2/5B mRNA; Lanes 2–4: same with 10, 20 and 40 pmol, respectively, of aIF2/5B mRNA. Bottom. Curve obtained by quantifying with a Molecular Dynamics Phosphoimager the radioactivity retained in the bands as shown in the top panel. The relative increment on the -axis is the ratio between radioactivity values in the absence of added aIF2/5B mRNA and values measured in the presence of the indicated amounts of aIF2/5B mRNA

Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon -4

<p><b>Copyright information:</b></p><p>Taken from "Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon "</p><p></p><p>Molecular Microbiology 2007;65(3):700-713.</p><p>Published online 01 Aug 2007</p><p>PMCID:PMC1976387.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p

Protection of fMet-tRNAi from alkaline hydrolysis in the presence of native and chimeric IF2 proteins

<p><b>Copyright information:</b></p><p>Taken from "Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon "</p><p></p><p>Molecular Microbiology 2007;65(3):700-713.</p><p>Published online 01 Aug 2007</p><p>PMCID:PMC1976387.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p> The extent of fMet-tRNAi protection was determined as described in . Closed circle: recombinant aIF2/5B; closed diamond; recombinant IF2; closed triangle: BaSu1 chimera; closed square: BaSu2 chimera; open triangle: SuBa chimera; open circle: no proteins added

Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon -8

<p><b>Copyright information:</b></p><p>Taken from "Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon "</p><p></p><p>Molecular Microbiology 2007;65(3):700-713.</p><p>Published online 01 Aug 2007</p><p>PMCID:PMC1976387.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p

Amidated route: estimates of NMNAT isozymes' physiological activity.

a<p>Values are calculated from the steady-state kinetic <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113939#pone.0113939.e002" target="_blank">equation (2</a>) described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113939#s2" target="_blank">Methods</a>, using the NMNAT isozyme activities measured at saturating substrates (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113939#pone-0113939-g003" target="_blank">Figure 3C</a>) as <i>V</i>_max values, the NMN and ATP levels (nmol/g, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113939#pone-0113939-g004" target="_blank">Figure 4</a>) as micromolar concentrations, and the <i>K</i>_m values from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113939#pone.0113939-Orsomando1" target="_blank">[18]</a>; nd, not detectable.</p><p>Amidated route: estimates of NMNAT isozymes' physiological activity.</p

Correlation between the physiological levels of products (NAD or NaAD) and substrates (NMN or NaMN) of the NMNAT-catalyzed reaction in different mouse tissues.

<p>Average values ± standard errors are those depicted in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113939#pone-0113939-g004" target="_blank">Figure 4</a>. The “AMIDATED” refers to grouped NAD and NMN levels in the indicated tissues; the group “DEAMIDATED” refers to levels of NaAD and NaMN (smaller circle) in the same tissues (not detailed). The linear relationship in different tissues between the steady-state levels of NAD and NaAD relative to NMN and NaMN, respectively, is highlighted. The “outlier” square symbol (▪) represents blood, where diverging values were observed compared to all other examined tissues.</p

Metabolic pathway of NAD biosynthesis in mammals.

<p>The two routes, “amidated” and “deamidated”, are highlighted. The different sources of the pyridine moiety for NAD synthesis are circled. The metabolites involved are: nicotinic acid (Na), nicotinamide (Nam), nicotinamide riboside (NR), quinolinic acid (Qa), nicotinate mononucleotide (NaMN), nicotinamide mononucleotide (NMN), nicotinate adenine dinucleotide (NaAD), and nicotinamide adenine dinucleotide (NAD). The enzymes involved are: QaPRT (EC 2.4.2.19), NaPRT (EC 2.4.2.11), NamPRT (EC 2.4.2.12), NRK (EC 2.7.1.22), NMNAT (EC 2.7.7.1), and NADS (EC 6.3.5.1).</p