Evidence for the monomeric nature of thymosins

Haritos, A.A.; Yialouris, P.P.; Heimer, E.; Félix, Arthur; Hannappel, Ewald; Rosemeyer, Michael A.

doi:10.1016/0014-5793(89)80547-2

Cited by 19 publications

(16 citation statements)

References 17 publications

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“…Human pTK was produced in E. coli and puri¢ed by the hot phenol extraction method developed by Evsta¢eva et al [20], which is based on the unique tendency of pTK to accumulate in an aqueous phase rather than in an organic phase, like all bacterial proteins. The protein was puri¢ed further by anion exchange chromatography to more than 99% purity and showed characteristic features reported previously [7,23,24]. Thus, in the course of gel-¢ltration chromatography, pTK eluted as a protein with an apparent molecular mass of 58.4 þ 0.7 kDa (not shown) ; the molecular mass calculated from its amino acid composition is 12 kDa.…”

Section: Resultssupporting

confidence: 63%

Amyloid fibrils from the mammalian protein prothymosin α

et al. 2002

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Mammalian prothymosin K K, a small (12 kDa) and extremely acidic protein (pI 3.5), is a member of the growing family of 'natively' unfolded proteins. We demonstrate that at low pH (V V3) and high concentrations, prothymosin K K is capable of forming regular elongated fibrils with flat ribbon structure 4^5 nm in height and 12^13 nm in width as judged from scanning force and electron microscopy. These aggregates induced a characteristic spectral shift of thioflavin T fluorescence and their circular dichroism spectra were indicative of significant L L-sheet content, suggesting formation of classical amyloid. Our findings indicate that natively unfolded proteins may have a general propensity to form amyloid fibrils under conditions inducing partially folded conformations. ß 2002 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.

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Section: Resultssupporting

confidence: 63%

Amyloid fibrils from the mammalian protein prothymosin α

et al. 2002

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“…1), both in reducing and non-reducing conditions. This data agrees with the experimental data for gel filtration which we [21] and others [22] have obtained, deducing that ProTa presents a strong aggregating tendency.…”

Section: Labeling Of Prota With "'Isupporting

confidence: 93%

Prothymosin α receptors on peripheral blood mononuclear cells

1994

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'251-Labeled prothymosin a (ProTa) was used to study the presence and characteristics of receptors for ProTa on human peripheral blood mononuclear cells (PBMC). The kinetics of '251-ProTa binding to PBMC was fast at 37"C, whilst it required 50 min to reach equilibrium at 4°C and room temperature. Analysis of steady state binding data by the method of Scatchard and by unlabeled ProTa competition experiments identified two binding sites with an apparent equilibrium dissociation constant of 216321 pM for the high-affinity receptor and of 11.421.1 nM for the low-affinity one; the sites per cell ranged from 1,479 to 1,519 and from 47,547 to 56,169, respectively. The kinetically derived equilibrium dissociation constant agreed with these data and showed no interaction between receptors.

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“…It appears surprising that a peptide of only 5 kDa molecular mass does not freely diffuse through nuclear pores. However, structural studies have implicated that thymosin β 4 is an elongated molecule (Zarbock et al, 1990;Czisch et al, 1993;Ballweber et al, 2002) and data from gel filtration experiments have shown that it migrates like a protein of considerably higher molecular mass (Haritos et al, 1989). Future experiments will aim to identify the cytoplasmic factors involved in its nuclear translocation and binding partners.…”

Section: Discussionmentioning

confidence: 99%

Nuclear localisation of the G-actin sequestering peptide thymosin β4

Huff

Rosorius

Otto

et al. 2004

Journal of Cell Science

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Thymosin β4 is regarded as the main G-actin sequestering peptide in the cytoplasm of mammalian cells. It is also thought to be involved in cellular events like cancerogenesis, apoptosis, angiogenesis, blood coagulation and wound healing. Thymosin β4 has been previously reported to localise intracellularly to the cytoplasm as detected by immunofluorescence. It can be selectively labelled at two of its glutamine-residues with fluorescent Oregon Green cadaverine using transglutaminase; however, this labelling does not interfere with its interaction with G-actin. Here we show that after microinjection into intact cells, fluorescently labelled thymosin β4 has a diffuse cytoplasmic and a pronounced nuclear staining. Enzymatic cleavage of fluorescently labelled thymosin β4 with AsnC-endoproteinase yielded two mono-labelled fragments of the peptide. After microinjection of these fragments, only the larger N-terminal fragment, containing the proposed actin-binding sequence exhibited nuclear localisation, whereas the smaller C-terminal fragment remained confined to the cytoplasm. We further showed that in digitonin permeabilised and extracted cells, fluorescent thymosin β4 was solely localised within the cytoplasm, whereas it was found concentrated within the cell nuclei after an additional Triton X100 extraction. Therefore, we conclude that thymosin β4 is specifically translocated into the cell nucleus by an active transport mechanism, requiring an unidentified soluble cytoplasmic factor. Our data furthermore suggest that this peptide may also serve as a G-actin sequestering peptide in the nucleus, although additional nuclear functions cannot be excluded.

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Evidence for the monomeric nature of thymosins

Cited by 19 publications

References 17 publications

Amyloid fibrils from the mammalian protein prothymosin α

Amyloid fibrils from the mammalian protein prothymosin α

Prothymosin α receptors on peripheral blood mononuclear cells

Nuclear localisation of the G-actin sequestering peptide thymosin β4

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