2020
DOI: 10.1016/j.bpj.2020.08.023 View full text |Buy / Rent full text
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Abstract: Differential scanning calorimetry (DSC) indicated that PDZ3 undergoes a peculiar thermal denaturation exhibiting two endothermic peaks due to the formation of reversible oligomers at high temperature (N↔I 6 ↔D). This contrasts sharply with the standard 2-state denaturation model observed for small, globular proteins. We performed an alanine scanning analysis by individually mutating three hydrophobic residues at the crystallographic oligomeric interface (Phe340, Leu342, Ile389) and one away from the interface … Show more

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“…The absence of oligomers for the FD3A chimera suggests that FD3A preserves its one‐domain compact form and uses a different denaturation mechanism. It has been described that the PDZ3 oligomer intermediates are formed by interactions of PDZ β‐sheets 62 . Therefore, in the case of FD3A, a stable domain–domain interface between PDZ3 and TrpCage could be possibly preserved, and this could block the association of β‐sheets necessary to form the oligomeric intermediates.…”
Section: Resultsmentioning
“…The PDZ2 domain from the ZO-2 protein showed a T m around 40-50 C with an obvious protein concentration dependency. 61 In one of the previous studies, the T m of PDZ3 was estimated at a surprisingly high temperature of 72.58 C, 62 and the explanation of such high thermostability was provided as follows: the PDZ3 partially thermally unfolded intermediates form aggregates under native conditions, and they were detected by native-state hydrogen exchange experiments. It implies that two processes could take place during the thermal unfolding of the PDZ3 domain.…”
Section: Thermal and Chemical Denaturation Of Fd3a And Fd4a Revealed Distinct Chimera Charactersmentioning
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“…Recently, deviations from the two-state unfolding model have further been observed in small globular proteins under near-physiological conditions (PSD95-PDZ3 [16][17][18], DEN4 ED3 [19], Che Y [20], and Cro repressor [21]). These proteins show two endothermic peaks in the DSC thermogram measured under a reversible condition without precipitation or chemical alteration [16][17][18][19]. The second peak is assigned to the presence of a high-temperature partially unfolded intermediate that oligomerizes reversibly during the thermal denaturation [16][17][18][19].…”
Section: Introductionmentioning
“…These proteins show two endothermic peaks in the DSC thermogram measured under a reversible condition without precipitation or chemical alteration [16][17][18][19]. The second peak is assigned to the presence of a high-temperature partially unfolded intermediate that oligomerizes reversibly during the thermal denaturation [16][17][18][19]. In previous studies, we showed that the high-temperature RO of PDZ3 and DEN4 ED3 are inhibited by single mutations by constructing two variants having a single mutation at their crystallographic interfaces that undergo a fully reversible two-state thermal denaturation [18][19].…”
Section: Introductionmentioning