A model for the aggregation of the acylphosphatase from Sulfolobus solfataricus in its native-like state

Bemporad, Francesco; Vannocci, Tommaso; Varela, Lorena; Azuaga, Ana I.; Chiti, Fabrizio

doi:10.1016/j.bbapap.2008.08.021

Cited by 22 publications

(54 citation statements)

References 36 publications

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“…On the other hand the same data show that Sso AcP remains soluble in phosphate buffer, independently of the presence of TFE in solution. The persistent solubility of Sso AcP in phosphate buffer upon the addition of TFE is due to the fact that phosphate acts as a competitive inhibitor of acylphosphatases (28) with a K d of 1.12 Ϯ 0.10 mM in the case of Sso AcP at pH 5.5 (12). The binding of the phosphate ion to Sso AcP causes a dramatic increase in conformational stability and structural rigidity of the native state, inhibiting aggregation, which by contrast requires structural fluctuations within the native state (6).…”

Section: Mg MLmentioning

confidence: 99%

“…A combination of limited proteolysis experiments (10), protein engineering (10,12,13), and specific kinetic tests with peptides resembling the N-terminal segment (12) have put forward a model in which the rate-determining step for the first aggregation phase consists in the collision of the N-terminal segment with the globular unit of Sso AcP, probably with ␤-strand 4 (12). By contrast, the further conversion of the native-like assemblies into protofibrils appears to involve a cooperative and positionindependent conversion of the protein molecules present in the initial aggregates (10).…”

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confidence: 99%

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Structural and Dynamics Characteristics of Acylphosphatase from Sulfolobus solfataricus in the Monomeric State and in the Initial Native-like Aggregates

Pagano

Bemporad

Fogolari

et al. 2010

Journal of Biological Chemistry

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It has previously been shown that the acylphosphatase from Sulfolobus solfataricus is capable of forming amyloid-like aggregates under conditions in which the native structure is maintained and via the transient formation of native-like aggregates. Based on the previously determined NMR structure of the native protein, showing a ferredoxin-like fold and the peculiar presence of an unstructured N-terminal segment, we show here, at a molecular level using NMR spectroscopy, that indeed S. solfataricus acylphosphatase remains in a native-like conformation when placed in aggregating conditions and that such a nativelike structure persists when the protein forms the initial aggregates, at least within the low molecular weight species. The analysis carried out under different solution conditions, based on the measurement of the combined 1 H and 15 N chemical shifts and hydrogen/deuterium exchange rates, enabled the most significant conformational changes to be monitored upon transfer of the monomeric state into aggregating conditions and upon formation of the initial native-like aggregates. Important increases of the hydrogen/deuterium exchange rates throughout the native protein, accompanied by small and localized structural changes, in the monomeric protein were observed. The results also allow the identification of the intermolecular interaction regions within the native-like aggregates, that involve, in particular, the N-terminal unstructured segment, the apical region including strands S4 and S5 with the connecting loop, and the opposite active site.Proteins and peptides have a generic propensity to form well organized aggregates characterized by a fibrillar morphology and an extended cross-␤ structure, generally referred to as amyloid-like fibrils (1, 2). This process is important for a number of reasons. From a physicochemical perspective, it represents an essential feature of the behavior of polypeptide chains that need to be fully understood for a thorough characterization of the nature of proteins (3). From a more biological perspective, formation of amyloid fibrils, or intracellular inclusions with structurally related characteristics, is associated with over 40 pathological conditions in humans (4, 5). It is also a major problem in biotechnology because the large scale expression of proteins potentially interesting to the market often results in their self-assembly in inclusion bodies with amyloid-like structural features (4).

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Section: Mg MLmentioning

confidence: 99%

mentioning

confidence: 99%

Structural and Dynamics Characteristics of Acylphosphatase from Sulfolobus solfataricus in the Monomeric State and in the Initial Native-like Aggregates

Pagano

Bemporad

Fogolari

et al. 2010

Journal of Biological Chemistry

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“…Although the process of native-like aggregation is well characterized in vitro for Sso AcP (21,22,(24)(25)(26)(27)(28)(29)(30) and for several other proteins (11,15,20), similar processes in vivo have not yet been described. In this study, we addressed this issue by expressing in Escherichia coli wild-type (WT) Sso AcP and three of its mutants, which are devoid of the N-terminal segment, contain a destabilizing mutation in the globular unit, or display both modifications.…”

Section: Introductionmentioning

confidence: 99%

“…The intermolecular interactions responsible for the first phase have been shown to involve the interaction of the edge b-strand 4 (labeled in Fig. 1 A) of one protein molecule with the N-terminal segment of a second molecule; the b-strand 4 of the second molecule can then interact with the N-terminal segment of a third molecule, leading to a polymerization cascade (21,(26)(27)(28). The b-strand 4 of Sso AcP appears to be protected only poorly against edge-to-edge interactions compared to other acylphosphatases (29).…”

Section: Introductionmentioning

confidence: 99%

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Direct Conversion of an Enzyme from Native-like to Amyloid-like Aggregates within Inclusion Bodies

Elia

Cantini

Chiti

et al. 2017

Biophysical Journal

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The acylphosphatase from Sulfolobus solfataricus (Sso AcP) is a globular protein able to aggregate in vitro from a native-like conformational ensemble without the need for a transition across the major unfolding energy barrier. This process leads to the formation of assemblies in which the protein retains its native-like structure, which subsequently convert into amyloid-like aggregates. Here, we investigate the mechanism by which Sso AcP aggregates in vivo to form bacterial inclusion bodies after expression in E. coli. Shortly after the initiation of expression, Sso AcP is incorporated into inclusion bodies as a native-like protein, still exhibiting small but significant enzymatic activity. Additional experiments revealed that this overall process of aggregation is enhanced by the presence of the unfolded N-terminal region of the sequence and by destabilization of the globular segment of the protein. At later times, the Sso AcP molecules in the inclusion bodies lose their native-like properties and convert into b-sheet-rich amyloid-like structures, as indicated by their ability to bind thioflavin T and Congo red. These results show that the aggregation behavior of this protein is similar in vivo to that observed in vitro, and that, at least for a predominant part of the protein population, the transition from a native to an amyloid-like structure occurs within the aggregate state.

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Amyloid Aggregation of Bacillus circulans Xylanase under Native Conditions and its Modulation by β‐Amyloid‐Derived Peptide Fragments

et al. 2018

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The aggregation of intrinsically disordered proteins into fibrils is implicated in many neurodegenerative diseases. Amyloid aggregation is a generic property of proteins as evidenced by globular proteins that often form amyloid aggregates under partially denaturing conditions. Recently, multiple lines of evidence have suggested that the amyloid aggregation of globular proteins can also occur under native conditions. Unfortunately, amyloid aggregation under native conditions has been demonstrated in only a handful of cases. Engineering a globular protein's amyloid aggregation might benefit from its fusion to an amyloid‐derived fragment with reduced aggregation propensity. Unfortunately, the impacts of such fragments on the amyloid aggregation under native conditions have yet to be examined. In this study, we show that a globular protein, Bacillus circulans xylanase (BCX), can aggregate to form amyloid fibrils under native conditions. When BCX was mixed with or fused to the non‐self‐aggregating fragments, KLVFWAK and ELVFWAE—which were derived from β‐amyloid (Aβ)—they modulated the BCX amyloid aggregation to differing extents. This study also provides insight into a correlation between the kinetic stability and amyloid aggregation of BCX, and supports a view that Aβ‐derived fragments can be useful for the modulating amyloid aggregation of some, though not all, proteins.

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A model for the aggregation of the acylphosphatase from Sulfolobus solfataricus in its native-like state

Cited by 22 publications

References 36 publications

Structural and Dynamics Characteristics of Acylphosphatase from Sulfolobus solfataricus in the Monomeric State and in the Initial Native-like Aggregates

Structural and Dynamics Characteristics of Acylphosphatase from Sulfolobus solfataricus in the Monomeric State and in the Initial Native-like Aggregates

Direct Conversion of an Enzyme from Native-like to Amyloid-like Aggregates within Inclusion Bodies

Amyloid Aggregation of Bacillus circulans Xylanase under Native Conditions and its Modulation by β‐Amyloid‐Derived Peptide Fragments

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