9
Biosynthesis of Bacterial Cellulose
Genes.involved.in.the.synthesis.and.cleavage.of.c-di-GMP.were.first.identified. using. oligonucleotide. primers. designed. from. the. partial. amino. acid.
sequence. of. a. purified. diguanylate. cyclase. (Tal. et. al.. 1998).. These. primers.
were. used. to. synthesize. a. short. fragment. by. polymerase. chain. reaction.
(PCR),. and. through. screening. of. a. genomic. library. from. G. xylinus. strain.
1306-11.and.genetic.analysis.of.mutants,.three.operons.(cdg1,.cdg2,.cdg3).for.
turnover. of. c-di-GMP. were. identified. in. this. strain.. Each. operon. carries. a.
diguanylate. cyclase. (dgc). gene. for. synthesis. of. c-di-GMP. and. a. phosphodiesterase-A.(pdeA).gene.for.cleavage.of.c-di-GMP..The.diguanylate.cyclase.
and.phosphodiesterase. proteins.coded.for.in.these.operons. were. found.to.
have.the.GGDEF.and.EAL.sequence.motifs,.and.these.motifs.have.since.been.
identified.in.a.large.number.of.proteins.in.bacteria..Proteins.containing.these.
motifs.are.known.to.influence.a.variety.of.activities.through.synthesis.and.
breakdown.of.c-di-GMP,.which.is.now.recognized.as.an.important.signaling.
molecule.in.bacteria.
What is Known About the Proteins Involved
in Cellulose Synthesis in Bacteria?
So.far,.analyses.of.proteins.encoded.by.genes.in.the.bcs.operon.and.by.genes.
present. adjacent. to. this. operon. in. G. xylinus. have. provided. only. limited.
information.as.to.their.role.in.cellulose.biosynthesis..Some.of.these.proteins.
have.been.identified.biochemically.and.a.few.of.their.features.determined.
experimentally,.while.for.others.their.known.features.are.based.on.sequence.
analysis. and. structure. determination.. These. proteins. are. discussed. here.
briefly.to.highlight.what.is.currently.known.about.them.and.their.possible.
role.in.cellulose.synthesis.in.G. xylinus.
CMCase.is.a.secreted.protein.coded.by.ORF1.(cmcax).that.exhibits.endoβ-1,4-glucanase.activity..The.structure.of.this.protein.has.been.determined.
and. it. belongs. to. the. glycoside. hydrolase. family. 8. (Yasutake. et. al.. 2006)..
Expression.of.the.cmcax.gene.is.induced.by.gentiobiose.in.the.medium,.and.
the.presence.of.CMCase.protein.in.the.culture.medium.results.in.an.increase.
in.cellulose.production..It.is.not.very.clear.as.to.what.role.this.protein.may.
play.in.cellulose.production,.but.it.has.been.suggested.to.be.partly.attached.
to.the.cell.surface.where.it.affects.organization.of.the.glucan.chains.in.the.
assembly.of.the.cellulose.ribbon.
The.role.of.the.protein.coded.by.ORF2.(ccp).was.identified.through.characterization. of. Cel − . mutants. of. G. xylinus.. Not. much. is. known. about. this.
protein,.although.it.has.been.suggested.to.have.a.role.in.both.the.production.
and.crystallization.of.cellulose.I.(Nakai.et.al..2002).
Biosynthesis of Bacterial Cellulose
Genes.involved.in.the.synthesis.and.cleavage.of.c-di-GMP.were.first.identified. using. oligonucleotide. primers. designed. from. the. partial. amino. acid.
sequence. of. a. purified. diguanylate. cyclase. (Tal. et. al.. 1998).. These. primers.
were. used. to. synthesize. a. short. fragment. by. polymerase. chain. reaction.
(PCR),. and. through. screening. of. a. genomic. library. from. G. xylinus. strain.
1306-11.and.genetic.analysis.of.mutants,.three.operons.(cdg1,.cdg2,.cdg3).for.
turnover. of. c-di-GMP. were. identified. in. this. strain.. Each. operon. carries. a.
diguanylate. cyclase. (dgc). gene. for. synthesis. of. c-di-GMP. and. a. phosphodiesterase-A.(pdeA).gene.for.cleavage.of.c-di-GMP..The.diguanylate.cyclase.
and.phosphodiesterase. proteins.coded.for.in.these.operons. were. found.to.
have.the.GGDEF.and.EAL.sequence.motifs,.and.these.motifs.have.since.been.
identified.in.a.large.number.of.proteins.in.bacteria..Proteins.containing.these.
motifs.are.known.to.influence.a.variety.of.activities.through.synthesis.and.
breakdown.of.c-di-GMP,.which.is.now.recognized.as.an.important.signaling.
molecule.in.bacteria.
What is Known About the Proteins Involved
in Cellulose Synthesis in Bacteria?
So.far,.analyses.of.proteins.encoded.by.genes.in.the.bcs.operon.and.by.genes.
present. adjacent. to. this. operon. in. G. xylinus. have. provided. only. limited.
information.as.to.their.role.in.cellulose.biosynthesis..Some.of.these.proteins.
have.been.identified.biochemically.and.a.few.of.their.features.determined.
experimentally,.while.for.others.their.known.features.are.based.on.sequence.
analysis. and. structure. determination.. These. proteins. are. discussed. here.
briefly.to.highlight.what.is.currently.known.about.them.and.their.possible.
role.in.cellulose.synthesis.in.G. xylinus.
CMCase.is.a.secreted.protein.coded.by.ORF1.(cmcax).that.exhibits.endoβ-1,4-glucanase.activity..The.structure.of.this.protein.has.been.determined.
and. it. belongs. to. the. glycoside. hydrolase. family. 8. (Yasutake. et. al.. 2006)..
Expression.of.the.cmcax.gene.is.induced.by.gentiobiose.in.the.medium,.and.
the.presence.of.CMCase.protein.in.the.culture.medium.results.in.an.increase.
in.cellulose.production..It.is.not.very.clear.as.to.what.role.this.protein.may.
play.in.cellulose.production,.but.it.has.been.suggested.to.be.partly.attached.
to.the.cell.surface.where.it.affects.organization.of.the.glucan.chains.in.the.
assembly.of.the.cellulose.ribbon.
The.role.of.the.protein.coded.by.ORF2.(ccp).was.identified.through.characterization. of. Cel − . mutants. of. G. xylinus.. Not. much. is. known. about. this.
protein,.although.it.has.been.suggested.to.have.a.role.in.both.the.production.
and.crystallization.of.cellulose.I.(Nakai.et.al..2002).