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Enhancing human spermine synthase activity by engineered mutations.

Zhang Z, Zheng Y, Petukh M, Pegg A, Ikeguchi Y, Alexov E - PLoS Comput. Biol. (2013)

Bottom Line: It is confirmed experimentally that the variant, bearing four amino acid substitutions, is catalytically more active than the wild type.The study demonstrates that wild type SMS is not particularly evolutionarily optimized with respect to the reaction spermidine → spermine.Having in mind that currently there are no variations (non-synonymous single nucleotide polymorphism, nsSNP) detected in healthy individuals, it can be speculated that the human SMS function is precisely tuned toward its wild type and any deviation is unwanted and disease-causing.

View Article: PubMed Central - PubMed

Affiliation: Computational Biophysics and Bioinformatics, Department of Physics, Clemson University, Clemson, South Carolina, United States of America.

ABSTRACT
Spermine synthase (SMS) is an enzyme which function is to convert spermidine into spermine. It was shown that gene defects resulting in amino acid changes of the wild type SMS cause Snyder-Robinson syndrome, which is a mild-to-moderate mental disability associated with osteoporosis, facial asymmetry, thin habitus, hypotonia, and a nonspecific movement disorder. These disease-causing missense mutations were demonstrated, both in silico and in vitro, to affect the wild type function of SMS by either destabilizing the SMS dimer/monomer or directly affecting the hydrogen bond network of the active site of SMS. In contrast to these studies, here we report an artificial engineering of a more efficient SMS variant by transferring sequence information from another organism. It is confirmed experimentally that the variant, bearing four amino acid substitutions, is catalytically more active than the wild type. The increased functionality is attributed to enhanced monomer stability, lowering the pKa of proton donor catalytic residue, optimized spatial distribution of the electrostatic potential around the SMS with respect to substrates, and increase of the frequency of mechanical vibration of the clefts presumed to be the gates toward the active sites. The study demonstrates that wild type SMS is not particularly evolutionarily optimized with respect to the reaction spermidine → spermine. Having in mind that currently there are no variations (non-synonymous single nucleotide polymorphism, nsSNP) detected in healthy individuals, it can be speculated that the human SMS function is precisely tuned toward its wild type and any deviation is unwanted and disease-causing.

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Potential distribution.(A) Electrostatic field lines for the WT HsSMS; Drawing method: FieldLines; GardientMag: 1.45; Min Length: 35.31; Max Length: 50.90; Coloring Scale Data Range: −1; 1; (B) Potential difference (mutant – WT) mapped onto HsSMS surface; Coloring method: Volume; 1. Drawing method: Surf; Coloring Scale Data Range: −1; 1 (blue – positive, red – negative).
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pcbi-1002924-g002: Potential distribution.(A) Electrostatic field lines for the WT HsSMS; Drawing method: FieldLines; GardientMag: 1.45; Min Length: 35.31; Max Length: 50.90; Coloring Scale Data Range: −1; 1; (B) Potential difference (mutant – WT) mapped onto HsSMS surface; Coloring method: Volume; 1. Drawing method: Surf; Coloring Scale Data Range: −1; 1 (blue – positive, red – negative).

Mentions: One plausible reason why HsSMS functions as a dimer could be that the dimerization is necessary to provide guidance and steer the positively charged SPD/SPM in and out the active site. It can be speculated that there are two paths shown in Figure 2A as path “A” and “B”. The electrostatic field lines form a kind of funnel leading to the vicinity of the active sites. The path “A” is in the cleft between HsSMS domains and is a single path. In contrast, the path “B” is perpendicular to the dimer axis and is symmetrical (doubled) with an angle of symmetry 180° (Figure 2A). Neither of these paths, “A” or “B”, exist if one uses in the modeling the HsSMS monomer only, i.e. they are product of the dimerization. The mutations further increase the magnitude of the electrostatic potential along the both paths as it is illustrated in Figure 2B, where the change of the potential due to the mutations is shown. The potential difference is mapped onto molecular surface of the dimer. Red color patch indicates that the potential is more negative in the dimer as compared with the WT, and blue color the opposite. It can be seen that patches corresponding to both paths, “A” and “B”, are more negative in the mutant than in the WT, providing support of the hypothesis that the mutant enhanced activity is caused by better steering of the substrates to the active site.


Enhancing human spermine synthase activity by engineered mutations.

Zhang Z, Zheng Y, Petukh M, Pegg A, Ikeguchi Y, Alexov E - PLoS Comput. Biol. (2013)

Potential distribution.(A) Electrostatic field lines for the WT HsSMS; Drawing method: FieldLines; GardientMag: 1.45; Min Length: 35.31; Max Length: 50.90; Coloring Scale Data Range: −1; 1; (B) Potential difference (mutant – WT) mapped onto HsSMS surface; Coloring method: Volume; 1. Drawing method: Surf; Coloring Scale Data Range: −1; 1 (blue – positive, red – negative).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3585406&req=5

pcbi-1002924-g002: Potential distribution.(A) Electrostatic field lines for the WT HsSMS; Drawing method: FieldLines; GardientMag: 1.45; Min Length: 35.31; Max Length: 50.90; Coloring Scale Data Range: −1; 1; (B) Potential difference (mutant – WT) mapped onto HsSMS surface; Coloring method: Volume; 1. Drawing method: Surf; Coloring Scale Data Range: −1; 1 (blue – positive, red – negative).
Mentions: One plausible reason why HsSMS functions as a dimer could be that the dimerization is necessary to provide guidance and steer the positively charged SPD/SPM in and out the active site. It can be speculated that there are two paths shown in Figure 2A as path “A” and “B”. The electrostatic field lines form a kind of funnel leading to the vicinity of the active sites. The path “A” is in the cleft between HsSMS domains and is a single path. In contrast, the path “B” is perpendicular to the dimer axis and is symmetrical (doubled) with an angle of symmetry 180° (Figure 2A). Neither of these paths, “A” or “B”, exist if one uses in the modeling the HsSMS monomer only, i.e. they are product of the dimerization. The mutations further increase the magnitude of the electrostatic potential along the both paths as it is illustrated in Figure 2B, where the change of the potential due to the mutations is shown. The potential difference is mapped onto molecular surface of the dimer. Red color patch indicates that the potential is more negative in the dimer as compared with the WT, and blue color the opposite. It can be seen that patches corresponding to both paths, “A” and “B”, are more negative in the mutant than in the WT, providing support of the hypothesis that the mutant enhanced activity is caused by better steering of the substrates to the active site.

Bottom Line: It is confirmed experimentally that the variant, bearing four amino acid substitutions, is catalytically more active than the wild type.The study demonstrates that wild type SMS is not particularly evolutionarily optimized with respect to the reaction spermidine → spermine.Having in mind that currently there are no variations (non-synonymous single nucleotide polymorphism, nsSNP) detected in healthy individuals, it can be speculated that the human SMS function is precisely tuned toward its wild type and any deviation is unwanted and disease-causing.

View Article: PubMed Central - PubMed

Affiliation: Computational Biophysics and Bioinformatics, Department of Physics, Clemson University, Clemson, South Carolina, United States of America.

ABSTRACT
Spermine synthase (SMS) is an enzyme which function is to convert spermidine into spermine. It was shown that gene defects resulting in amino acid changes of the wild type SMS cause Snyder-Robinson syndrome, which is a mild-to-moderate mental disability associated with osteoporosis, facial asymmetry, thin habitus, hypotonia, and a nonspecific movement disorder. These disease-causing missense mutations were demonstrated, both in silico and in vitro, to affect the wild type function of SMS by either destabilizing the SMS dimer/monomer or directly affecting the hydrogen bond network of the active site of SMS. In contrast to these studies, here we report an artificial engineering of a more efficient SMS variant by transferring sequence information from another organism. It is confirmed experimentally that the variant, bearing four amino acid substitutions, is catalytically more active than the wild type. The increased functionality is attributed to enhanced monomer stability, lowering the pKa of proton donor catalytic residue, optimized spatial distribution of the electrostatic potential around the SMS with respect to substrates, and increase of the frequency of mechanical vibration of the clefts presumed to be the gates toward the active sites. The study demonstrates that wild type SMS is not particularly evolutionarily optimized with respect to the reaction spermidine → spermine. Having in mind that currently there are no variations (non-synonymous single nucleotide polymorphism, nsSNP) detected in healthy individuals, it can be speculated that the human SMS function is precisely tuned toward its wild type and any deviation is unwanted and disease-causing.

Show MeSH
Related in: MedlinePlus