1eyd: Difference between revisions

Revision as of 13:32, 21 February 2008

STRUCTURE OF WILD-TYPE S. NUCLEASE AT 1.7 A RESOLUTION

OverviewOverview

Seven hyper-stable multiple mutants have been constructed in staphylococcal nuclease by various combinations of eight different stabilizing single mutants. The stabilities of these multiple mutants determined by guanidine hydrochloride denaturation were 3.4 to 5.6 kcal/mol higher than that of the wild-type. Their thermal denaturation midpoint temperatures were 12.6 to 22.9 deg. C higher than that of the wild-type. These are among the greatest increases in protein stability and thermal denaturation midpoint temperature relative to the wild-type yet attained. There has been great interest in understanding how proteins found in thermophilic organisms are stabilized. One frequently cited theory is that the packing of hydrophobic side-chains is improved in the cores of proteins isolated from thermophiles when compared to proteins from mesophiles. The crystal structures of four single and five multiple stabilizing mutants of staphylococcal nuclease were solved to high resolution. No large overall structural change was found, with most changes localized around the sites of mutation. Rearrangements were observed in the packing of side-chains in the major hydrophobic core, although none of the mutations was in the core. It is surprising that detailed structural analysis showed that packing had improved, with the volume of the mutant protein's hydrophobic cores decreasing as protein stability increased. Further, the number of van der Waals interactions in the entire protein showed an experimentally significant increase correlated with increasing stability. These results indicate that optimization of packing follows as a natural consequence of increased protein thermostability and that good packing is not necessarily the proximate cause of high stability. Another popular theory is that thermostable proteins have more electrostatic and hydrogen bonding interactions and these are responsible for the high stabilities. The mutants here show that increased numbers of electrostatic and hydrogen bonding interactions are not obligatory for large increases in protein stability.

About this StructureAbout this Structure

1EYD is a Single protein structure of sequence from Staphylococcus aureus. Active as Micrococcal nuclease, with EC number 3.1.31.1 Full crystallographic information is available from OCA.

ReferenceReference

Increasing the thermostability of staphylococcal nuclease: implications for the origin of protein thermostability., Chen J, Lu Z, Sakon J, Stites WE, J Mol Biol. 2000 Oct 20;303(2):125-30. PMID:11023780

Page seeded by OCA on Thu Feb 21 12:32:50 2008

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-[[Image:1eyd.jpg|left|200px]]<br /><applet load="1eyd" size="450" color="white" frame="true" align="right" spinBox="true"
+[[Image:1eyd.jpg|left|200px]]<br /><applet load="1eyd" size="350" color="white" frame="true" align="right" spinBox="true"
 caption="1eyd, resolution 1.70&Aring;" />
 '''STRUCTURE OF WILD-TYPE S. NUCLEASE AT 1.7 A RESOLUTION'''<br />
 ==Overview==
-Seven hyper-stable multiple mutants have been constructed in, staphylococcal nuclease by various combinations of eight different, stabilizing single mutants. The stabilities of these multiple mutants, determined by guanidine hydrochloride denaturation were 3.4 to 5.6, kcal/mol higher than that of the wild-type. Their thermal denaturation, midpoint temperatures were 12.6 to 22.9 deg. C higher than that of the, wild-type. These are among the greatest increases in protein stability and, thermal denaturation midpoint temperature relative to the wild-type yet, attained. There has been great interest in understanding how proteins, found in thermophilic organisms are stabilized. One frequently cited, theory is that the packing of hydrophobic side-chains is improved in the, cores of proteins isolated from thermophiles when compared to proteins, from mesophiles. The crystal structures of four single and five multiple, stabilizing mutants of staphylococcal nuclease were solved to high, resolution. No large overall structural change was found, with most, changes localized around the sites of mutation. Rearrangements were, observed in the packing of side-chains in the major hydrophobic core, although none of the mutations was in the core. It is surprising that, detailed structural analysis showed that packing had improved, with the, volume of the mutant protein's hydrophobic cores decreasing as protein, stability increased. Further, the number of van der Waals interactions in, the entire protein showed an experimentally significant increase, correlated with increasing stability. These results indicate that, optimization of packing follows as a natural consequence of increased, protein thermostability and that good packing is not necessarily the, proximate cause of high stability. Another popular theory is that, thermostable proteins have more electrostatic and hydrogen bonding, interactions and these are responsible for the high stabilities. The, mutants here show that increased numbers of electrostatic and hydrogen, bonding interactions are not obligatory for large increases in protein, stability.
+Seven hyper-stable multiple mutants have been constructed in staphylococcal nuclease by various combinations of eight different stabilizing single mutants. The stabilities of these multiple mutants determined by guanidine hydrochloride denaturation were 3.4 to 5.6 kcal/mol higher than that of the wild-type. Their thermal denaturation midpoint temperatures were 12.6 to 22.9 deg. C higher than that of the wild-type. These are among the greatest increases in protein stability and thermal denaturation midpoint temperature relative to the wild-type yet attained. There has been great interest in understanding how proteins found in thermophilic organisms are stabilized. One frequently cited theory is that the packing of hydrophobic side-chains is improved in the cores of proteins isolated from thermophiles when compared to proteins from mesophiles. The crystal structures of four single and five multiple stabilizing mutants of staphylococcal nuclease were solved to high resolution. No large overall structural change was found, with most changes localized around the sites of mutation. Rearrangements were observed in the packing of side-chains in the major hydrophobic core, although none of the mutations was in the core. It is surprising that detailed structural analysis showed that packing had improved, with the volume of the mutant protein's hydrophobic cores decreasing as protein stability increased. Further, the number of van der Waals interactions in the entire protein showed an experimentally significant increase correlated with increasing stability. These results indicate that optimization of packing follows as a natural consequence of increased protein thermostability and that good packing is not necessarily the proximate cause of high stability. Another popular theory is that thermostable proteins have more electrostatic and hydrogen bonding interactions and these are responsible for the high stabilities. The mutants here show that increased numbers of electrostatic and hydrogen bonding interactions are not obligatory for large increases in protein stability.
 ==About this Structure==
-EYD is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Staphylococcus_aureus Staphylococcus aureus]. Active as [http://en.wikipedia.org/wiki/Micrococcal_nuclease Micrococcal nuclease], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.1.31.1 3.1.31.1] Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1EYD OCA].
+EYD is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Staphylococcus_aureus Staphylococcus aureus]. Active as [http://en.wikipedia.org/wiki/Micrococcal_nuclease Micrococcal nuclease], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.1.31.1 3.1.31.1] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1EYD OCA].
 ==Reference==
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 [[Category: Lu, Z.]]
 [[Category: Sakon, J.]]
-[[Category: Stites, W.E.]]
+[[Category: Stites, W E.]]
 [[Category: hydrolase]]
-''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Tue Nov 20 14:25:37 2007''
+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 12:32:50 2008''