Chemistry Reference
In-Depth Information
the.amino.acid.can.change.depending.on.its.location.and.surrounding.amino.acids..
The.second.shell.coordination.characteristics.identify.which.residues.are.the.most.
and.least.hydrated.and.therefore.the.most.and.least.polar,.respectively,.within.that.
speciic. protein.. The. second. shell. coordination. values. are. obtained. by. taking. the.
difference. between. the. coordination. number. at. 7. and. 4.Å.. The. second. shell. coor-
dination. numbers. for. these. proteins. exhibit. different. values. than. in. the. irst. shell..
The. least. coordinated. residues. (with. a. coordination. number. of. ∼18). in. the. second.
shell.are.two.glycine.residues,.25.and.51,.in.the.N-.and.C-helices.of.the.wild-type.
protein..The.most.hydrated.residue.is.again.the.C-terminal.alanine,.with.a.coordina-
tion.of.34.water.molecules..In.the.case.of.the.mutant-type.protein,.the.least.hydrated.
residues.are.the.residue.9.cysteine.and.the.residue.47.glycine.(∼16.coordinated.water.
molecules).and.the.most.hydrated.is.again.the.C-terminal.alanine.(∼33.coordinated.
water.molecules)..The.differences.between.the.wild-type.and.the.mutant.are.not.only.
evident.in.the.most.and.least.hydrated.residues,.but.the.A53T.mutant.has.on.average.
a.lower.coordination.number.throughout.the.protein..This.difference.further.empha-
sizes.that.the.presence.of.aqueous.solution.in.the.irst.and.second.coordination.shells.
can.yield.insight.into.the.in.vivo.behavior.of.the.protein.and.that.the.impact.of.water.
cannot.be.ignored.
In.some.experimental.techniques,.such.as.electrospray.ionization.mass.spectrom-
etry.(ESI-MS),.the.impact.of.water.is.not.observed.because.the.protein.is.converted.
to.an.ionized.gaseous.state.for.a.short.time.before.it.enters.the.solution.and.is.ana-
lyzed.. Several. papers. have. been. published. on. the. use. of. ESI-MS. to. characterize.
the.structure.of.proteins..In.a.1993.paper.by.Mirza.et.al.,.they.described.a.method.
of. studying. conformational. changes. in. bovine. ubiquitin. and. bovine. cytochrome. c.
by. ESI-MS. using. heat. to. denature. the. proteins. from. their. natively. folded. state. as.
they. pass. through. the. ESI. needle. into. the. mass. spectrometer.
68
. Studies. have. also.
been. performed. by. Kaitashov. and. Eyles. on. biomolecular. conformations. and. con-
formational.dynamics.of.protein.folding.via.ESI-MS.
69
.A.recent.article.by.Ashcroft.
describes. ESI-MS. studies. coupled. with. ion. mobility. spectroscopy. to. study. ibril.
formation.by.amyloids.
11
.While.these.techniques.do.provide.information.about.the.
intermolecular.(in.the.case.of.multiple.protein.systems).and.intramolecular.hydrogen.
bonding. for. proteins,. the. impact. of. water. on. these. structures. cannot. be. observed.
because.the.measurements.occur.while.the.proteins.are.in.the.gas.phase.
11
.Although.
techniques.are.available.to.denature.proteins.by.heat.or.pH,.the.structure.observed.
may.still.differ.from.the.native.in.vivo.conformation..According.to.these.papers,.the.
ESI-MS.and.native.structures.of.the.proteins.were.found.to.be.comparable;.however,.
the.proteins.studied.had.highly.folded.structures.in.vitro.
68,69
.Proteins.that.are.highly.
folded. may. not. have. drastic. structural. changes. from. aqueous. solution. to. the. gas.
phase,.but.in.the.case.of.natively.unfolded.proteins.such.as.synucleins,.the.structures.
are.very.different,.as.shown.earlier..Mass.spectrometry.techniques.have.been.very.
useful.in.determining.the.structure.of.proteins.and.their.aggregates.in.the.gas.phase.
and. also. discerning. the. relative. amounts. of. the. components,. in. the. case. of. aggre-
gates,. that.are.present..However,. in.order.to.understand.the.mechanism.of.in.vivo.
aggregation,.the.impact.of.water.should.be.considered.because.it.can.have.a.signii-
cant.effect.on.the.formation.of.aggregates..Transporting.a.highly.folded.gas.phase.
structure.into.solution.may.not.result.in.reformation.of.the.wild-type.structure.since.
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