Post on 13-Feb-2016
description
Archaeal extreme halophiles(halobacteria, haloarchaea)
require high concentration of NaCl for growth
found in Great Salt Lake, Dead Sea, evaporation ponds
bacteriorhodopsin – light-activated proton pump – covalently attached retinal – red color
1 family, 34 genera
heterotrophs
mostly aerobic
Adaptation to high salt
high internal concentration of KCl
halorhodopsin – light-activatedchloride pump, pumps chloride in
protein adaptations to high salt
excess acidic amino acids
fewer hydrophobic amino acids
Halophiles with sequenced genomes
Halobacterium salinarum NRC-1 and R1
Haloarcula marismortui
Haloquadratum walsbyi
Natronomonas pharaonis
Haloferax volcanii
New genomes in our study
Halorubrum lacusprofundi - CSP
Halogeometricum borinquense - GEBA
Halomicrobium mukohataei - GEBA
Halorhabdus utahensis - GEBA
Haloterrigena turkmenica – GEBA
Isolation Grows on
Halobacterium salted cow hide complex medium or amino acids
Haloarcula Dead Sea water Glc, Frc, Scr, Gly, Ac, Suc, Mal (Rib, Xyl)
Haloquadratum saltern water, Spain, Australia Pyruvate, DHA
Natronomonas desert lake water, Egypt Pyr, Fum, Form, But
Haloferax Dead Sea sediment Yeast extract, tryptone, Glc, Xyl, Gly, Ac, Lac
Halorubrum Deep Lake, Antarctica Glc, Gal, Man, Rib, Lct, Gly, Suc, Lac, Ac, Prop, Eth, Form
Halogeometricum saltern water, Puerto Rico Glc, Frc, Man, Xyl, Mlt, Tre, Cel, Raf, Gly (Ara, Rib, Lac, Scr)
Halomicrobium salt flat soil, Argentina Glc, Gal, Scr, Mlt, Gly (Man, Rib)
Halorhabdus Great Salt Lake sediment Glc, Xyl, Frc, Xylan
Haloterrigena saline soil, Turkmenia Glc, Frc, Man, Scr, Mlt
Questions
● What genes are conserved in all ten genomes?
● What are the haloarchaeal signature genes (in all haloarchaea but not in other archaea)?
● What are the organism-specific genes in each genome?
● Have some genomes recently lost genes?
● Are there genes found in water halophiles but not in soil/sediment halophiles?
● Are there genes found in soil/sediment halophiles but not in water halophiles?
All of these questions can be answered using the phylogenetic profiler.
Genes conserved in all halophiles (core genes)
Genes conserved in all halophiles (core genes)
40% of total genes in most genomes
50% of total genes in Halobacterium NRC-1
This % of conserved genes is about the same as other families with multiple genomes from different genera
Genes conserved in all halophiles (core genes)
Part of gene list from phylogenetic profiler results.
Core genes - results
Transcription, Translation, DNA replication
RNA processing – sRNPs, RNAse P, methyltransferases
Protein processing – chaperones, proteases
Protein secretion – signal-dependent secretion, twin arginine
Signal transduction – kinases, phosphatases, response regulators
Amino acid synthesis – most, but not Leu, Ile, Val, Lys
Purine/pyrimidine synthesis – except guanylate kinase, thymidylate synthase
Cofactor synthesis – heme, pyridoxal, flavins, F420
Lipid synthesis – mevalonate pathway, phospholipid metabolism
Central metabolism – TCA cycle, most of glycolysis
Haloarchaeal signature – genes present in all haloarchaea but not in other archaea
First, select all Archaea
Second, select halophiles (I also removed draft genomes)
Haloarchaeal signature genes
Haloarchaeal signature genes
Most have unknown functions (even the ones with COGs)
but a few are involved in gamma-glutamylcysteine metabolism
Central metabolism in the IMG network
Pentose phosphate pathway (PPP)
● haloarchaea are thought to use the oxidative PPP
● produces pentoses for RNA, DNA synthesis
● produces NADPH for biosynthesis of amino acids, etc.
Add the IMG terms to the function cart, do a profile with the ten haloarchaea.
key enzyme is missing in Halorhabdus utahensis
Check COGs also
Some archaea use the reverse ribulose monophosphate pathway
The two key enzymes are not in any haloarchaea.
There is one more pathway – nonoxidative PPP
fructose-6P hexulose-6P
ribulose-5P+
formaldehyde
Function profile – nonoxidative PPP
Halorhabdus has all the enzymes required for nonoxidative PPP.
Why does it have a different pathway?
fructose-6P+
glyceraldehyde-3P
erythrose-4P+
xylulose-5P
fructose-6P+
glyceraldehyde-3P+
sedoheptulose-7P
ribose-5P+
xylulose-5P
Different pathways of D-xylose catabolism
D-xylose
D-xylulose
D-xylulose-5P
xylose isomerase, Huta_2443
xylulokinase, Huta_2446
non-oxidative pentose phosphate pathway
D-xylose
D-xylonolactone
D-xylonate
2-keto-3-deoxyxylonate
2-ketoglutarate semialdehyde
2-ketoglutarateHalorhabdus
Halorubrum, Haloferax, Haloterrigena, Haloarcula
L-arabinose metabolism
L-arabinose
L-ribuloseL-arabinose isomerase, Huta_1154
L-ribulose-5Pribulokinase, Huta_1150?
D-xylulose-5Pribulose-5P 4-epimerase, Huta_1149
non-oxidative pentose phosphate pathway
not in any other archaea
only in Halorhabdus
Conclusion:
Non-oxidative PPP is reversible, may make it easier to grow on pentoses.
How does Halorhabdus generate NADPH?
In other halophiles, oxidative PPP produces NADPH
NADP+ + NADH NAD+ + NADPH
Halorhabdus is the only haloarchaeon to have NAD/NADP transhydrogenase.
glycerol glycerol-3P DHAPATP ADP quinone quinol
glycolysis
Glycerol metabolism and transport
Glycerol is an important carbon/energy source for halophiles. All except N. pharaonis have glycerol kinase and glycerol-3P DHase
But only Halomicrobium mukohataei has an identified glycerol transporter
glycerol-3P DHase
glycerol kinase
predicted membrane
protein
glycerol transporter, MIP family
Hbor has 2 glycerol kinases, both adj to conserved membrane protein
In two bacteria, the same membrane protein is adjacent to glycerol kinase
glycerol kinase
In two bacteria, the membrane protein is adjacent to 1,3-propanediol dehydrogenase
1,3-propanediol dehydrogenase
In five bacteria, the membrane protein is close to glycerol dehydratase
small subunit large subunit
We have used a not well characterized group of organisms to predict gene function in model organisms (E. coli, Rhodobacter)
Motility in haloarchaea
Flagellar proteins, chemotaxis proteins
Gas vesicle proteins
H. walsbyi lacks flagellar motility but has gas vesicle proteins
Sediment halophiles lack gas vesicle proteins
Haloterrigena turkmenica was isolated from soil.
Why does it have gas vesicle proteins?
Isolated from upper crust of solonchak, poorly drained soil
keep haloarchaea close to light and oxygen