Sandbox Reserved 325: Difference between revisions

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=Chorismate Mutase=

==Introduction==

The gene Rv1885c from ''Mycobacteriam tuberculosis'' encodes for a non-functional chorismate mutase (*MtCM)<ref name="pizza" />. This non-functional mutase has a 33-amino-acid cleavable sequence <ref name="pizza" />. It is a vital enzyme in the shikimate pathway, which allows for the synthesis of tryptophan, tyrosinem and phenylalanine <ref name="pizza" />. Chorismate mutase provides a 2x10<sup>6</sup> fold increase in the rate of reacrion in comparision to the uncatalyzed reaction <ref > P.D. Lyne, A.J. Mulholland, W.G. Richards. Insights into chorismate mutase catalysis from a combined qm/mm simulation of the enzyme reaction. Journal of the American Chemistry Society. 1995 117(45):11345-11350</ref>. Chorismate mutase only occurs in bacteria, higher plants, and fungi, due to the fact that the shikimate pathway is only found in these organisms <ref name="strat" />. In ''Escherichia coli'', chorismate mutase has a periplasmic destination<ref name="pizza" />. In ''M. tuberculosis'' there is in abscence of a periplasmic compartment for chorismate mutase, so it secretes into the culture filtrate of ''M. tuberculosis''<ref name="pizza" />. It is believed that a pseudoperiplasmic space might exist in ''M. tuberculosis''<ref name="pizza" />. Rv1885c is synthesized along with the 33-amino-acid terminal sequence, which when expressed with ''Escherichia coli'', is cleaved off the mature protein<ref name="pizza" />. Chorismate mutase is the only example of an enzyme catalyzing a percyclic reaction <ref name="strat"> PMID:10960481 </ref>

The gene Rv1885c from ''Mycobacteriam tuberculosis'' encodes for a non-functional chorismate mutase (*MtCM)<ref name="pizza" />. This non-functional mutase has a 33-amino-acid cleavable sequence <ref name="pizza" />. It is a vital enzyme in the shikimate pathway, which allows for the synthesis of tryptophan, tyrosinem and phenylalanine <ref name="pizza" />. This protein acts at the first branch point of the shikimate pathway, making it a regulating step<ref name="CMW2"> PMID:PMC55368 </ref>. Chorismate mutase provides a 2x10<sup>6</sup> fold increase in the rate of reacrion in comparision to the uncatalyzed reaction <ref > P.D. Lyne, A.J. Mulholland, W.G. Richards. Insights into chorismate mutase catalysis from a combined qm/mm simulation of the enzyme reaction. Journal of the American Chemistry Society. 1995 117(45):11345-11350</ref>. Chorismate mutase only occurs in bacteria, higher plants, and fungi, due to the fact that the shikimate pathway is only found in these organisms <ref name="strat" />. In ''Escherichia coli'', chorismate mutase has a periplasmic destination<ref name="pizza" />. In ''M. tuberculosis'' there is in abscence of a periplasmic compartment for chorismate mutase, so it secretes into the culture filtrate of ''M. tuberculosis''<ref name="pizza" />. It is believed that a pseudoperiplasmic space might exist in ''M. tuberculosis''<ref name="pizza" />. Rv1885c is synthesized along with the 33-amino-acid terminal sequence, which when expressed with ''Escherichia coli'', is cleaved off the mature protein<ref name="pizza" />. Chorismate mutase is the only example of an enzyme catalyzing a percyclic reaction <ref name="strat"> PMID:10960481 </ref>

Chorismate mutase acts at the first branch-point of aromatic amino

acid biosynthesis and catalyzes the conversion of chorismate to

prephenate. WIKI 2

We also demonstrate that unlike the corresponding

proteins of E. coli, Mtb chorismate mutase

does not have any associated prephenate dehydratase

or dehydrogenase activity, indicating its monofunctional

nature 22222222222

The

predicted N-terminal signal sequence of Mtb chorismate

mutase was capable of functioning as one in E. coli,

suggesting that Mtb chorismate mutase belongs to the

AroQ class of chorismate mutases. It was evident that

Rv1885c may not be the only enzyme with chorismate

mutase enzyme function within Mtb, based on our observation

of the presence of chorismate mutase activity2222222222222222

Mycobacterium tuberculosis (Mtb)1 has developed ingenious

mechanisms to survive inside the hostile environment presented

by the host and to acquire essential nutrients from this

adverse environment (1–3). The emergence of drug-resistant

strains and synergy with the AIDS virus has further aggravated

the disease scenario (4–6). For the development of new

therapeutic intervention strategies, there is a need for identification

of novel targets that are not only unique to Mtb but

blocking of which would either prove lethal to the bacterium or

render it extremely susceptible to the host immune response.

In this context, understanding the mechanism of action of the

aromatic amino acid pathway enzymes of Mtb assumes the

utmost importance because most of the corresponding genes

have been proven essential for the bacterium and have no

human or mammalian counterpart (7, 8). Moreover, amino acid

auxotrophs of Mtb do not survive or multiply in macrophages

(9, 10), suggesting that these amino acids are not available

within the compartment of the macrophage in which the bacteria

reside.

displayed by another hypothetical protein coded by

open reading frame Rv0948c, a novel instance of the

existence of two monofunctional chorismate mutase 2222222222222222222222222

==Structure==

Line 27:

Line 69:

ph tolerance from 4.0 to 7.5 for optimal activity

active site exists though hydrogen bonding and electrostatic interactions with chorismate

no beta sheets 222222

Mtb Chorismate Mutase Is a Dimeric Protein with a Predominantly

�-Helical Structure—Whereas catalytic activity and

regulatory activity of Mtb chorismate mutase point toward

some novel properties of the enzyme, the study was continued

to determine the biophysical parameters of the enzyme to define

the actual class to which it belongs. Size exclusion chromatography

was performed to determine the oligomeric state of

the protein. The output was a single peak corresponding to the

dimeric state of the recombinant protein (data not shown). In this

context, Mtb chorismate mutase is similar to the E. coli or yeast

chorismate mutases, which are also dimers of identical subunits

(24, 36). To determine the secondary structure of Mtb chorismate

mutase, the CD spectrum was recorded on a JASON spectropolarimeter

(Fig. 6). The data were analyzed using the K2D software

available on-line. The results suggest a predominantly

�-helical structure for the enzyme. This is reminiscent of the

AroQ class of enzymes from yeast and E. coli (24, 37) that are also

helical proteins. Members of the AroQ class of chorismate mutases

consist of unregulated and regulated (AroQr) enzymes and

are unusually divergent among closely related organisms (38).

This structure showed 71% helices with essentially no �-sheets. 2222222222

Sandbox Reserved 325: Difference between revisions

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@@ Line 4: / Line 4: @@
 =Chorismate Mutase=
 ==Introduction==
-The gene Rv1885c from ''Mycobacteriam tuberculosis'' encodes for a non-functional chorismate mutase (*MtCM)<ref name="pizza" />.  This non-functional mutase has a 33-amino-acid cleavable sequence <ref name="pizza" />.  It is a vital enzyme in the shikimate pathway, which allows for the synthesis of tryptophan, tyrosinem and phenylalanine <ref name="pizza" />.  Chorismate mutase provides a 2x10<sup>6</sup> fold increase in the rate of reacrion in comparision to the uncatalyzed reaction <ref > P.D. Lyne, A.J. Mulholland, W.G. Richards. Insights into chorismate mutase catalysis from a combined qm/mm simulation of the enzyme reaction. Journal of the American Chemistry Society. 1995 117(45):11345-11350</ref>.  Chorismate mutase only occurs in bacteria, higher plants, and fungi, due to the fact that the shikimate pathway is only found in these organisms <ref name="strat" />.  In ''Escherichia coli'', chorismate mutase has a periplasmic destination<ref name="pizza" />.  In ''M. tuberculosis'' there is in abscence of a periplasmic compartment for chorismate mutase, so it secretes into the culture filtrate of ''M. tuberculosis''<ref name="pizza" />.  It is believed that a pseudoperiplasmic space might exist in ''M. tuberculosis''<ref name="pizza" />. Rv1885c is synthesized along with the 33-amino-acid terminal sequence, which when expressed with ''Escherichia coli'', is cleaved off the mature protein<ref name="pizza"  />.  Chorismate mutase is the only example of an enzyme catalyzing a percyclic reaction <ref name="strat"> PMID:10960481 </ref>
+The gene Rv1885c from ''Mycobacteriam tuberculosis'' encodes for a non-functional chorismate mutase (*MtCM)<ref name="pizza" />.  This non-functional mutase has a 33-amino-acid cleavable sequence <ref name="pizza" />.  It is a vital enzyme in the shikimate pathway, which allows for the synthesis of tryptophan, tyrosinem and phenylalanine <ref name="pizza" />.  This protein acts at the first branch point of the shikimate pathway, making it a regulating step<ref name="CMW2"> PMID:PMC55368 </ref>.  Chorismate mutase provides a 2x10<sup>6</sup> fold increase in the rate of reacrion in comparision to the uncatalyzed reaction <ref > P.D. Lyne, A.J. Mulholland, W.G. Richards. Insights into chorismate mutase catalysis from a combined qm/mm simulation of the enzyme reaction. Journal of the American Chemistry Society. 1995 117(45):11345-11350</ref>.  Chorismate mutase only occurs in bacteria, higher plants, and fungi, due to the fact that the shikimate pathway is only found in these organisms <ref name="strat" />.  In ''Escherichia coli'', chorismate mutase has a periplasmic destination<ref name="pizza" />.  In ''M. tuberculosis'' there is in abscence of a periplasmic compartment for chorismate mutase, so it secretes into the culture filtrate of ''M. tuberculosis''<ref name="pizza" />.  It is believed that a pseudoperiplasmic space might exist in ''M. tuberculosis''<ref name="pizza" />. Rv1885c is synthesized along with the 33-amino-acid terminal sequence, which when expressed with ''Escherichia coli'', is cleaved off the mature protein<ref name="pizza"  />.  Chorismate mutase is the only example of an enzyme catalyzing a percyclic reaction <ref name="strat"> PMID:10960481 </ref>
+Chorismate mutase acts at the first branch-point of aromatic amino
+acid biosynthesis and catalyzes the conversion of chorismate to
+prephenate. WIKI 2
+We also demonstrate that unlike the corresponding
+proteins of E. coli, Mtb chorismate mutase
+does not have any associated prephenate dehydratase
+or dehydrogenase activity, indicating its monofunctional
+nature 22222222222
+The
+predicted N-terminal signal sequence of Mtb chorismate
+mutase was capable of functioning as one in E. coli,
+suggesting that Mtb chorismate mutase belongs to the
+AroQ class of chorismate mutases. It was evident that
+Rv1885c may not be the only enzyme with chorismate
+mutase enzyme function within Mtb, based on our observation
+of the presence of chorismate mutase activity2222222222222222
+Mycobacterium tuberculosis (Mtb)1 has developed ingenious
+mechanisms to survive inside the hostile environment presented
+by the host and to acquire essential nutrients from this
+adverse environment (1–3). The emergence of drug-resistant
+strains and synergy with the AIDS virus has further aggravated
+the disease scenario (4–6). For the development of new
+therapeutic intervention strategies, there is a need for identification
+of novel targets that are not only unique to Mtb but
+blocking of which would either prove lethal to the bacterium or
+render it extremely susceptible to the host immune response.
+In this context, understanding the mechanism of action of the
+aromatic amino acid pathway enzymes of Mtb assumes the
+utmost importance because most of the corresponding genes
+have been proven essential for the bacterium and have no
+human or mammalian counterpart (7, 8). Moreover, amino acid
+auxotrophs of Mtb do not survive or multiply in macrophages
+(9, 10), suggesting that these amino acids are not available
+within the compartment of the macrophage in which the bacteria
+reside.
+displayed by another hypothetical protein coded by
+open reading frame Rv0948c, a novel instance of the
+existence of two monofunctional chorismate mutase 2222222222222222222222222
 ==Structure==
@@ Line 27: / Line 69: @@
 ph tolerance from 4.0 to 7.5 for optimal activity
 active site exists though hydrogen bonding and electrostatic interactions with chorismate
+no beta sheets 222222
+Mtb Chorismate Mutase Is a Dimeric Protein with a Predominantly
+�-Helical Structure—Whereas catalytic activity and
+regulatory activity of Mtb chorismate mutase point toward
+some novel properties of the enzyme, the study was continued
+to determine the biophysical parameters of the enzyme to define
+the actual class to which it belongs. Size exclusion chromatography
+was performed to determine the oligomeric state of
+the protein. The output was a single peak corresponding to the
+dimeric state of the recombinant protein (data not shown). In this
+context, Mtb chorismate mutase is similar to the E. coli or yeast
+chorismate mutases, which are also dimers of identical subunits
+(24, 36). To determine the secondary structure of Mtb chorismate
+mutase, the CD spectrum was recorded on a JASON spectropolarimeter
+(Fig. 6). The data were analyzed using the K2D software
+available on-line. The results suggest a predominantly
+�-helical structure for the enzyme. This is reminiscent of the
+AroQ class of enzymes from yeast and E. coli (24, 37) that are also
+helical proteins. Members of the AroQ class of chorismate mutases
+consist of unregulated and regulated (AroQr) enzymes and
+are unusually divergent among closely related organisms (38).
+This structure showed 71% helices with essentially no �-sheets. 2222222222