An interface point‐mutation variant of triosephosphate isomerase is compactly folded and monomeric at low protein concentrations

Borchert, Torben V.; Zeelen, J.P.; Schliebs, Wolfgang; Callens, Mia; Minke, Wendy E.; Jaenicke, Rainer; Wierenga, Rik K.

doi:10.1016/0014-5793(95)00586-x

Cited by 30 publications

(30 citation statements)

References 18 publications

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“…[41][42][43][44][45] These attempts to monomerize TIM involved deletions in the interfacial loop 3 and mutations that reversed charge pairing. We hypothesized that by choosing the most common amino acid at each position of cTIM, we had scrambled necessary amino acid interactions (i.e., correlations) at the dimer interface.…”

Section: Engineering the Interface Of Ctimmentioning

confidence: 99%

“…The structure of TIM suggests that dimerization is necessary for full assembly of the active site by the interdigitation of loop 3 from the opposite monomer, and engineered monomeric TIMs exhibit k cat /K m values reduced by about 10 4 -fold. [41][42][43][44][45] The quaternary structure of cTIM was determined by gel-filtration chromatography (Fig. 1c).…”

Section: Consensus Timmentioning

confidence: 99%

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Triosephosphate Isomerase by Consensus Design: Dramatic Differences in Physical Properties and Activity of Related Variants

Sullivan

Durani

Magliery

2011

Journal of Molecular Biology

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Section: Engineering the Interface Of Ctimmentioning

confidence: 99%

Section: Consensus Timmentioning

confidence: 99%

Triosephosphate Isomerase by Consensus Design: Dramatic Differences in Physical Properties and Activity of Related Variants

Sullivan

Durani

Magliery

2011

Journal of Molecular Biology

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“…Several studies on triosephosphate isomerases from various sources showed that different mutations in the dimer interface lead to monomers with different levels of activity. Some of these mutants showed concentration-dependent specific activity, suggesting that the inactive monomers can associate, at least to some degree, to give an active dimer (4,19,27,34).…”

mentioning

confidence: 99%

“…By using such an approach, it has been shown that mutations of different enzymes resulted in either active or inactive monomers (1,24,26). One of the most studied enzymes in this regard is triosephosphate isomerase, which usually forms a stable and tightly bound dimer (4). Several studies on triosephosphate isomerases from various sources showed that different mutations in the dimer interface lead to monomers with different levels of activity.…”

mentioning

confidence: 99%

Effect of Dimer Dissociation on Activity and Thermostability of the α-Glucuronidase fromGeobacillus stearothermophilus: Dissecting the Different Oligomeric Forms of Family 67 Glycoside Hydrolases

et al. 2004

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The oligomeric organization of enzymes plays an important role in many biological processes, such as allosteric regulation, conformational stability and thermal stability. ␣-Glucuronidases are family 67 glycosidases that cleave the ␣-1,2-glycosidic bond between 4-O-methyl-D-glucuronic acid and xylose units as part of an array of hemicellulose-hydrolyzing enzymes. Currently, two crystal structures of ␣-glucuronidases are available, those from Geobacillus stearothermophilus (AguA) and from Cellvibrio japonicus (GlcA67A). Both enzymes are homodimeric, but surprisingly their dimeric organization is different, raising questions regarding the significance of dimerization for the enzymes' activity and stability. Structural comparison of the two enzymes suggests several elements that are responsible for the different dimerization organization. Phylogenetic analysis shows that the ␣-glucuronidases AguA and GlcA67A can be classified into two distinct subfamilies of bacterial ␣-glucuronidases, where the dimer-forming residues of each enzyme are conserved only within its own subfamily. It seems that the different dimeric forms of AguA and GlcA67A represent the two alternative dimeric organizations of these subfamilies. To study the biological significance of the dimerization in ␣-glucuronidases, we have constructed a monomeric form of AguA by mutating three of its interface residues (W328E, R329T, and R665N). The activity of the monomer was significantly lower than the activity of the wild-type dimeric AguA, and the optimal temperature for activity of the monomer was around 35°C, compared to 65°C of the wild-type enzyme. Nevertheless, the melting temperature of the monomeric protein, 72.9°C, was almost identical to that of the wild-type, 73.4°C. It appears that the dimerization of AguA is essential for efficient catalysis and that the dissociation into monomers results in subtle conformational changes in the structure which indirectly influence the active site region and reduce the activity. Structural and mechanistic explanations for these effects are discussed.

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“…That is, in the intermediate state, the compact domain of the chain has substantially less structural fluctuations than the non-compact portion. Goldberg, 1980Banik et al, 1992Herold & Kirschner, 1990Mei et al, 1992Aceto et al, 1992Akasaka et al, 1982Shaw et al, 1995Borchert et al, 1995 "The 3-state dimers. The table describes the PDB code with the chain names, name of the molecule, the resolution of the structure, number of amino acids in a single chain, and the reference which shows the dimerization is 3-state.…”

Section: Dimer Kineticsmentioning

confidence: 99%

Mechanism and evolution of protein dimerization

Tsai

Nussinov

1998

Protein Science

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We have investigated the mechanism and the evolutionary pathway of protein dimerization through analysis of experimental structures of dimers. We propose that the evolution of dimers may have multiple pathways, including (1) formation of a functional dimer directly without going through an ancestor monomer, (2) formation of a stable monomer as an intermediate followed by mutations of its surface residues, and (3). a domain swapping mechanism, replacing one segment in a monomer by an equivalent segment from an identical chain in the dimer. Some of the dimers which are governed by a domain swapping mechanism may have evolved at an earlier stage of evolution via the second mechanism. Here, we follow the theory that the kinetic pathway reflects the evolutionary pathway. We analyze the structure-kinetics-evolution relationship for a collection of symmetric homodimers classified into three groups: (1) 14 dimers, which were referred to as domain swapping dimers in the literature; (2) nine 2-state dimers, which have no measurable intermediates in equilibrium denaturation; and (3), eight 3-state dimers, which have stable intermediates in equilibrium denaturation. The analysis consists of the following stages: (i) The dimer is divided into two structural units, which have twofold symmetry.,Each unit contains a contiguous segment from one polypeptide chain of the dimer, and its complementary contiguous segment from the other chain. (ii) The division is repeated progressively, with different combinations of the two segments in each unit. (iii) The coefficient of compactness is calculated for the units in all divisions. The coefficients obtained for different cuttings of a dimer form a compactness profile. The profile probes the structural organization of the two chains in a dimer and the stability of the monomeric state. We describe the features of the compactness profiles in each of the three dimer groups. The profiles identify the swapping segments in domain swapping dimers, and can usually predict whether a dimer has domain swapping. The kinetics of dimerization indicates that some dimers which have been assigned in the literature as domain swapping cases, dimerize through the 2-state kinetics, rather than through swapping segments of performed monomers. The compactness profiles indicate a wide spectrum in the kinetics of dimerization: dimers having no intermediate stable monomers; dimers having an intermediate with a stable monomer structure; and dimers having an intermediate with a stable structure in part of the monomer. These correspond to the multiple evolutionary pathways for dimer formation. The evolutionary mechanisms proposed here for dimers are applicable to other oligomers as well.Keywords: compactness; dimerization; domain swapping; intermediate; kinetics; oligomer evolution Oligomers are often the functional forms of proteins. It has been a challenging problem in protein science to understand the mechanism of oligomerization and the evolution of protein oligomers. The importance of this problem is re...

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An interface point‐mutation variant of triosephosphate isomerase is compactly folded and monomeric at low protein concentrations

Cited by 30 publications

References 18 publications

Triosephosphate Isomerase by Consensus Design: Dramatic Differences in Physical Properties and Activity of Related Variants

Triosephosphate Isomerase by Consensus Design: Dramatic Differences in Physical Properties and Activity of Related Variants

Effect of Dimer Dissociation on Activity and Thermostability of the α-Glucuronidase fromGeobacillus stearothermophilus: Dissecting the Different Oligomeric Forms of Family 67 Glycoside Hydrolases

Mechanism and evolution of protein dimerization

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