Differential scanning calorimetry as a tool for protein folding and stability

Johnson, Christopher M.

doi:10.1016/j.abb.2012.09.008

Cited by 317 publications

(207 citation statements)

References 99 publications

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“…Further heating, on the other hand, causes transition to conformations that become structurally trapped when cooled. Such a phenomenon has been observed in several other proteins (47,48). The estimated melting point of approximately 85°C for the minor capsid protein is in line with the melting temperatures of the major capsid proteins (49), demonstrating the extreme heat stability of all P23-77 virus capsid-associated proteins characterized so far.…”

Section: Vp11 Sequence Analysissupporting

confidence: 50%

The Minor Capsid Protein VP11 of Thermophilic Bacteriophage P23-77 Facilitates Virus Assembly by Using Lipid-Protein Interactions

Pawlowski

Moilanen

Rissanen

et al. 2015

J Virol

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Thermus thermophilus bacteriophage P23-77 is the type member of a new virus family of icosahedral, tailless, inner-membranecontaining double-stranded DNA (dsDNA) viruses infecting thermophilic bacteria and halophilic archaea. The viruses have a unique capsid architecture consisting of two major capsid proteins assembled in various building blocks. We analyzed the function of the minor capsid protein VP11, which is the third known capsid component in bacteriophage P23-77. Our findings show that VP11 is a dynamically elongated dimer with a predominantly ␣-helical secondary structure and high thermal stability. The high proportion of basic amino acids in the protein enables electrostatic interaction with negatively charged molecules, including nucleic acid and large unilamellar lipid vesicles (LUVs). The plausible biological function of VP11 is elucidated by demonstrating the interactions of VP11 with Thermus-derived LUVs and with the major capsid proteins by means of the dynamic-lightscattering technique. In particular, the major capsid protein VP17 was able to link VP11-complexed LUVs into larger particles, whereas the other P23-77 major capsid protein, VP16, was unable to link VP11-comlexed LUVs. Our results rule out a previously suggested penton function for VP11. Instead, the electrostatic membrane association of VP11 triggers the binding of the major capsid protein VP17, thus facilitating a controlled incorporation of the two different major protein species into the assembling capsid. IMPORTANCEThe study of thermophilic viruses with inner membranes provides valuable insights into the mechanisms used for stabilization and assembly of protein-lipid systems at high temperatures. Our results reveal a novel way by which an internal membrane and outer capsid shell are linked in a virus that uses two different major protein species for capsid assembly. We show that a positive protein charge is important in order to form electrostatic interactions with the lipid surface, thereby facilitating the incorporation of other capsid proteins on the membrane surface. This implies an alternative function for basic proteins present in the virions of other lipid-containing thermophilic viruses, whose proposed role in genome packaging is based on their capability to bind DNA. The unique minor capsid protein of bacteriophage P23-77 resembles in its characteristics the scaffolding proteins of tailed phages, though it constitutes a substantial part of the mature virion. V iruses from extremely hot environments need to develop special strategies to cope with high temperature. Hence, they provide a valuable source for the detection of novel enzymes for biotechnology and industrial processes. Their relative simple structure makes them an ideal model system for studying life at high temperatures. With respect to extreme thermophilic bacteria, the virus studied best on the structural level is P23-77, a strictly lytic phage infecting Thermus thermophilus ATCC 33923 (1, 2, 3). P23-77 is a representative of the novel Sphaerolipoviri...

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Section: Vp11 Sequence Analysissupporting

confidence: 50%

The Minor Capsid Protein VP11 of Thermophilic Bacteriophage P23-77 Facilitates Virus Assembly by Using Lipid-Protein Interactions

Pawlowski

Moilanen

Rissanen

et al. 2015

J Virol

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“…Figure 4 shows the DSC data collected for both the labeled and unlabeled mAb, carried out at a loading concentration of 0.42 mg/mL. The heat capacity profile for the unlabeled mAb looks quite similar to what has been seen in the literature for the IgG subclass (17). The shoulder at about 80°C corresponds to the melting of the C H 3 region within the Fc portion of the mAb, while the rather broad peak at 75.6°C corresponds to the F AB portion of the mAb.…”

Section: Sedimentation Velocitymentioning

confidence: 54%

Biophysical characterization of a model antibody drug conjugate

Arakawa

Kurosawa²,

Storms³

et al. 2016

DD&T

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“…Heaters around the cells, to keep the difference between cells equal to zero, respond increasing the temperature in the reference cell when the process is exothermic and increasing the temperature of the sample cell when the process is endothermic. The amount of energy necessary to maintain thermal balance within the system is proportional to the energy change occurring in the sample (PIRES et al, 2009;JOHNSON, 2013).…”

Section: δS (T) = δH (T T ) / T T -δC P Ln(t T |T) (3) δG (T) = δH (Tmentioning

confidence: 99%

“…DSC studies transitions or processes that gain or lose heat as a function of temperature in other words, when a substance is subjected to a temperature change, endothermic (heat absorption) or exothermic (heat generation) processes may occur. As most biological molecules of interest undergo transformations when subjected to temperature variations, it is possible to use DSC to determine the energy involved in such processes (JOHNSON, 2013).…”

Section: Differential Scanning Calorimetry (Dsc)mentioning

confidence: 99%