1auw: Difference between revisions

Revision as of 12:48, 21 February 2008

H91N DELTA 2 CRYSTALLIN FROM DUCK

OverviewOverview

The major soluble protein component of avian and reptilian eye lenses, delta crystallin, is highly homologous to the urea cycle enzyme, argininosuccinate lyase (ASL). In duck lenses there are two highly homologous delta crystallins, termed delta I and delta II, that are 94% identical in amino acid sequence. While delta II crystallin has been shown to exhibit ASL activity in vitro, delta I crystallin is inactive. The X-ray structure of a His to Asn mutant of duck delta II crystallin (H91N) has been determined to 2.5 A resolution using the molecular replacement technique. The overall fold of the protein is similar to other members of the superfamily to which this protein belongs, with the active site located in a cleft between three different monomers of the tetrameric protein. A reexamination of the kinetic properties of the H91N mutant reveals that the mutant has 10% wild-type activity. The Vmax of the mutant protein is identical to that of the wild-type protein, but a 10-fold increase in the Michaelis constant is seen, suggesting that His 91 is involved in binding the substrate. In an effort to determine the reasons for the loss of enzymatic activity in delta I crystallin, a structural comparison of the H91N mutant with the enzymatically inactive turkey delta I crystallin has been performed. This study revealed a remarkable similarity in the overall structures of the two proteins. Three regions of secondary structure do differ significantly between the two models; these include the N-terminal tail, a loop containing residues 76-91, and a cis versus trans peptide linkage at residue Thr 322. The cis to trans peptide variation appears to be an interspecies difference between turkey and duck and is therefore not directly involved in the loss of enzymatic activity. All the residues implicated in the catalytic mechanism are conserved in both the active and inactive proteins, and given the linearity of the relationship between the enzymatic activity of duck delta I/delta II heterotetramers and their delta II content (Piatigorsky & Horwitz, 1996), it is evident from the structure that only one of the three domains that contributes to the active site is responsible for the loss of activity in the delta I protein. Given the structural differences found in domain 1 (N-terminal tail and 76-91 loop), we postulate that these differences are responsible for the loss of catalytic activity in the delta I crystallin protein and that the delta I protein is inactive because it no longer binds the substrate.

About this StructureAbout this Structure

1AUW is a Single protein structure of sequence from Anas platyrhynchos. Active as Argininosuccinate lyase, with EC number 4.3.2.1 Known structural/functional Sites: , , and . Full crystallographic information is available from OCA.

ReferenceReference

Structural comparison of the enzymatically active and inactive forms of delta crystallin and the role of histidine 91., Abu-Abed M, Turner MA, Vallee F, Simpson A, Slingsby C, Howell PL, Biochemistry. 1997 Nov 18;36(46):14012-22. PMID:9369472

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 ==Overview==
-The major soluble protein component of avian and reptilian eye lenses, delta crystallin, is highly homologous to the urea cycle enzyme, argininosuccinate lyase (ASL). In duck lenses there are two highly, homologous delta crystallins, termed delta I and delta II, that are 94%, identical in amino acid sequence. While delta II crystallin has been shown, to exhibit ASL activity in vitro, delta I crystallin is inactive. The, X-ray structure of a His to Asn mutant of duck delta II crystallin (H91N), has been determined to 2.5 A resolution using the molecular replacement, technique. The overall fold of the protein is similar to other members of, the superfamily to which this protein belongs, with the active site, located in a cleft between three different monomers of the tetrameric, protein. A reexamination of the kinetic properties of the H91N mutant, reveals that the mutant has 10% wild-type activity. The Vmax of the mutant, protein is identical to that of the wild-type protein, but a 10-fold, increase in the Michaelis constant is seen, suggesting that His 91 is, involved in binding the substrate. In an effort to determine the reasons, for the loss of enzymatic activity in delta I crystallin, a structural, comparison of the H91N mutant with the enzymatically inactive turkey delta, I crystallin has been performed. This study revealed a remarkable, similarity in the overall structures of the two proteins. Three regions of, secondary structure do differ significantly between the two models; these, include the N-terminal tail, a loop containing residues 76-91, and a cis, versus trans peptide linkage at residue Thr 322. The cis to trans peptide, variation appears to be an interspecies difference between turkey and duck, and is therefore not directly involved in the loss of enzymatic activity., All the residues implicated in the catalytic mechanism are conserved in, both the active and inactive proteins, and given the linearity of the, relationship between the enzymatic activity of duck delta I/delta II, heterotetramers and their delta II content (Piatigorsky &amp; Horwitz, 1996), it is evident from the structure that only one of the three domains that, contributes to the active site is responsible for the loss of activity in, the delta I protein. Given the structural differences found in domain 1, (N-terminal tail and 76-91 loop), we postulate that these differences are, responsible for the loss of catalytic activity in the delta I crystallin, protein and that the delta I protein is inactive because it no longer, binds the substrate.
+The major soluble protein component of avian and reptilian eye lenses, delta crystallin, is highly homologous to the urea cycle enzyme, argininosuccinate lyase (ASL). In duck lenses there are two highly homologous delta crystallins, termed delta I and delta II, that are 94% identical in amino acid sequence. While delta II crystallin has been shown to exhibit ASL activity in vitro, delta I crystallin is inactive. The X-ray structure of a His to Asn mutant of duck delta II crystallin (H91N) has been determined to 2.5 A resolution using the molecular replacement technique. The overall fold of the protein is similar to other members of the superfamily to which this protein belongs, with the active site located in a cleft between three different monomers of the tetrameric protein. A reexamination of the kinetic properties of the H91N mutant reveals that the mutant has 10% wild-type activity. The Vmax of the mutant protein is identical to that of the wild-type protein, but a 10-fold increase in the Michaelis constant is seen, suggesting that His 91 is involved in binding the substrate. In an effort to determine the reasons for the loss of enzymatic activity in delta I crystallin, a structural comparison of the H91N mutant with the enzymatically inactive turkey delta I crystallin has been performed. This study revealed a remarkable similarity in the overall structures of the two proteins. Three regions of secondary structure do differ significantly between the two models; these include the N-terminal tail, a loop containing residues 76-91, and a cis versus trans peptide linkage at residue Thr 322. The cis to trans peptide variation appears to be an interspecies difference between turkey and duck and is therefore not directly involved in the loss of enzymatic activity. All the residues implicated in the catalytic mechanism are conserved in both the active and inactive proteins, and given the linearity of the relationship between the enzymatic activity of duck delta I/delta II heterotetramers and their delta II content (Piatigorsky &amp; Horwitz, 1996), it is evident from the structure that only one of the three domains that contributes to the active site is responsible for the loss of activity in the delta I protein. Given the structural differences found in domain 1 (N-terminal tail and 76-91 loop), we postulate that these differences are responsible for the loss of catalytic activity in the delta I crystallin protein and that the delta I protein is inactive because it no longer binds the substrate.
 ==About this Structure==
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 [[Category: Single protein]]
 [[Category: Abu-Abed, M.]]
-[[Category: Howell, P.L.]]
+[[Category: Howell, P L.]]
 [[Category: Vallee, F.]]
 [[Category: arginosuccinate lyase]]
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 [[Category: eye lens protein]]
-''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun Feb  3 09:31:40 2008''
+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 11:48:34 2008''