Karel Heremans

University of Leuven, Louvain, Flanders, Belgium

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Publications (100)245.88 Total impact

  • H. Pfeiffer, G. Klose, K. Heremans
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    ABSTRACT: Calorimetric scans of a number of low-hydrated phospholipids provide quite complex heat capacity profiles during the thermotropic main phase transition. This short study proposes some explanations that are based on experimental data obtained from four different methods (DSC, 31P-NMR, 2H-NMR, X-ray diffraction).The preliminary results show for POPC in which way DSC data are related to the melting of the hydrocarbon chains. Furthermore, the quadrupolar splitting data obtained from the 2H-NMR spectra of hydration water reveal how the water reorientation might contribute to the main phase transition of phospholipids.
    Chemical Physics Letters 05/2013; 572:120–124. · 2.15 Impact Factor
  • H Pfeiffer, G Klose, K Heremans
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    ABSTRACT: High-pressure investigations of phospholipids at low degrees of hydration are highly important due to the continuous interest in hydration related phenomena. This is for instance the case when discussing differences between the molecular behaviour under hydrostatic or hydration pressure, e.g. to characterise the structural and thermodynamic conditions that hold for the different theoretical approaches proposed in the literature. The stability of phospholipid aggregates in aqueous solutions is determined by a balance of attractive end repulsive forces as well as entropic and energetic interactions. The pressure-dependent variation of the frequency of molecular vibrations determined by FTIR spectroscopy might be an appropriate tool to study the nature of these interactions as well as of conformational changes for various degrees of hydrations. The paper reports preliminary results for the stretching and bending vibration of the CH2 groups that are mainly situated in the hydrocarbon chains. In a hydration range of about 2 water molecules per lipid in the case of DOPC and 6 water molecules for POPC, the pressure-dependent vibrations in the liquid crystalline phase change between red shift and blue shift. A further interesting parameter is the onset pressure for the correlation field splitting. It increases with dehydration, and it is assumed that the correlation field splitting does not only depend on the disorder related to the gauche conformer population in the hydrocarbon chains but also on the chain tilt in the gel phase.
    Chemistry and physics of lipids 03/2013; · 2.15 Impact Factor
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    ABSTRACT: It is well-known that fibrillogenesis of proteins can be influenced by diverse external parameters, such as temperature, pressure, agitation or chemical agents. The present preliminary study suggests that ultrasonic excitation at moderate intensities has a significant influence on the unfolding and aggregation behaviour of insulin. Irradiation with an average sound intensity of even as low as 70mW/cm(2) leads to a lowering of the unfolding and aggregation temperature up to 7K. The effect could be explained by an increase of the aggregation kinetics due to ultrasonically induced acoustic micro-streaming in the insulin solution that most probably enhances the aggregation rate. The clear and remarkable effect at relatively low sound intensities offers interesting options for further applications of ultrasound in biophysics and biochemistry. On the other hand, a process that causes a change of kinetics equivalent to 7K also gives a warning signal concerning the safety of those medical ultrasonic devices that work in this intensity range.
    Biochimica et Biophysica Acta 08/2012; · 4.66 Impact Factor
  • H Pfeiffer, H Weichert, G Klose, K Heremans
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    ABSTRACT: The hydration behaviour of mixtures of the zwitterionic phospholipid 1-palmitoyl-2-oleolyl-sn-glycero-3-phosphocholine (POPC) and the zwitterionic surfactant N,N-dimethyl-N-dodecyl-betain (C(12)-Bet) was investigated by sorption gravimetry, solid-state (31)P NMR-spectroscopy and small angle X-ray diffraction (SAXD). Negative excess hydration (dehydration) was found for almost all hydration degrees investigated. This behaviour is explained by the formation of an inner salt between the dipoles of phospholipid and surfactant headgroups that show a reverse sequence of partial charges with respect to the hydrocarbon backbone. The formation of an inner-salt most probably reduces potential water binding sites. Moreover, NMR data suggest that the incorporation of the zwitterionic surfactant into the phospholipid membrane is correlated with reorientation of the phosphate axis towards the membrane director as well as with reduced lateral and wobbling diffusion.
    Chemistry and physics of lipids 02/2012; 165(2):244-51. · 2.15 Impact Factor
  • Karel Heremans
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    ABSTRACT: This paper reviews some recent experimental observations on the behaviour of proteins under hydrostatic pressure. The results are correlated with findings on the influence of parameters such as temperature and pressure on living systems. Some processes are discussed which may be of importance for industrial applications such as high pressure food processing. In many cases a quantitative molecular interpretation is still lacking.
    07/2011: pages 443-469;
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    H Pfeiffer, G Klose, K Heremans
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    ABSTRACT: The hydration behaviour of equimolar mixtures of phospholipids and nonionic surfactants with different chain length was investigated by gravimetric sorption, NMR spectroscopy and X-ray diffraction. At the most hydration degrees investigated, the incorporation of nonionic surfactants in a phospholipid bilayer leads to an increase of the hydrophilicity, which can be shown by the presence of excess hydration. The increased hydrophilicity could be explained by the excavation of additional water binding sites due to the "dilution" of the dipole field of the phospholipid bilayer. Another related contribution arises from the increase of the accessible surface area due to the increase of gauche conformers that result from the steric mismatch when surfactants are incorporated into the phospholipid matrix. (2)H NMR spectroscopy was used to determine the quadrupolar splitting representing a measure of the order state of water. The swelling behaviour could be assessed by small-angle X-ray diffraction. (31)P NMR spectroscopy was applied for the assignment of phase structures to the respective hydration range.
    Chemistry and physics of lipids 03/2010; 163(3):318-28. · 2.15 Impact Factor
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    ABSTRACT: A variety of techniques, including high-pressure unfolding monitored by Fourier transform infrared spectroscopy, fluorescence, circular dichroism, and surface plasmon resonance spectroscopy, have been used to investigate the equilibrium folding properties of six single-domain antigen binders derived from camelid heavy-chain antibodies with specificities for lysozymes, β-lactamases, and a dye (RR6). Various denaturing conditions (guanidinium chloride, urea, temperature, and pressure) provided complementary and independent methods for characterizing the stability and unfolding properties of the antibody fragments. With all binders, complete recovery of the biological activity after renaturation demonstrates that chemical-induced unfolding is fully reversible. Furthermore, denaturation experiments followed by optical spectroscopic methods and affinity measurements indicate that the antibody fragments are unfolded cooperatively in a single transition. Thus, unfolding/refolding equilibrium proceeds via a simple two-state mechanism (N⇋U), where only the native and the denatured states are significantly populated. Thermally-induced denaturation, however, is not completely reversible, and the partial loss of binding capacity might be due, at least in part, to incorrect refolding of the long loops (CDRs), which are responsible for antigen recognition. Most interestingly, all the fragments are rather resistant to heat-induced denaturation (apparent Tm = 60–80°C), and display high conformational stabilities (ΔG(H2O) = 30–60 kJ mole−1). Such high thermodynamic stability has never been reported for any functional conventional antibody fragment, even when engineered antigen binders are considered. Hence, the reduced size, improved solubility, and higher stability of the camelid heavy-chain antibody fragments are of special interest for biotechnological and medical applications.
    Protein Science 04/2009; 11(3):500 - 515. · 2.74 Impact Factor
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    Helge Pfeiffer, Karel Heremans, Martine Wevers
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    ABSTRACT: a b s t r a c t This Letter reports a FTIR study on the pressure-induced unfolding of lysozyme in pure D 2 O close to the conditions of non excess hydration. An essential population of the proteins apparently refold into its native structure after pressure release. It could furthermore be shown that down to a hydration of h = 2 (h = m prot /m D 2 O ; m = mass), the unfolding pressure did not vary with hydration. Hydration depen-dent behaviour was found with respect to the change of the wavenumbers of the a-helical structure dur-ing unfolding. This result is discussed with respect to the reversibility of unfolding in pure solvents and with the effects of hydrogen exchange.
    Chemical Physics Letters 01/2009; 469(s 1–3). · 2.15 Impact Factor
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    ABSTRACT: FTIR spectroscopy was used to characterize and compare the temperature- and pressure-induced unfolding of ribonuclease A and a set of its variants engineered in a hydrophobic region of the C-terminal part of the molecule postulated as a CFIS. The results show for all the ribonucleases investigated, a cooperative, two-state, reversible unfolding transition using both pressure and temperature. The relative stabilities, among the different sites and different variants at the same site, monitored either through the changes in the position of the maximum of the amide I′ band and the tyrosine band, or the maximum of the band assigned to the β-sheet structure, corroborate the results of a previous study using fourth-derivative UV absorbance spectroscopy. In addition, variants at position 108 are the most critical for ribonuclease structure and stability. The V108G variant seems to present a greater conformational flexibility than the other variants. The pressure- and temperature-denaturated states of all the ribonucleases characterized retained some secondary structure. However, their spectral maxima were centered at different wavenumbers, which suggests that pressure- and temperature-denaturated states do not have the same structural characteristics. Nevertheless, there was close correlation between the pressure and temperature midpoint transition values for the whole series of protein variants, which indicated a common tendency of stability toward pressure and heat.
    Protein Science 12/2008; 10(4):725 - 734. · 2.74 Impact Factor
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    ABSTRACT: The stability of beta-glucosidase from the hyperthermophile Pyrococcus furiosus was studied as a function of pressure, temperature and pH. The conformational stability was monitored using FTIR spectroscopy, and the functional enzyme stability was monitored by inactivation studies. The enzyme proved to be highly piezostable and thermostable, with an unfolding pressure of 800 MPa at 85 degrees C. The tentative pressure-temperature stability diagram indicates that this enzyme is stabilized against thermal unfolding at low pressures. The activity measurements showed a two-step inactivation mechanism due to pressure that was most pronounced at lower temperatures. The first part of this inactivation took place at pressures below 300 MPa and was not visible as a conformational transition. The second transition in activity was concomitant with the conformational transition. An increase in pH from 5.5 to 6.5 was found to have a stabilizing effect.
    FEBS Journal 12/2008; 276(1):109-17. · 4.25 Impact Factor
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    Helge Pfeiffer, Karel Heremans, Martine Wevers
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    ABSTRACT: The elasticity of proteins, expressed by the compressibility, is potentially one of the most important properties of proteins because of the close relationship with its functionality. The compressibility of solutions can be determined by measurements of sound velocity and density. These quantities are related by the Newton-Laplace equation. In order to interpret the apparent compressibility of solutes in highly dilute solutions, it is required to consider the relation between compressibility and sound velocity of the solution using an appropriate reference system. The classical approach usually gives too small values for the apparent compressibility when compared with other methods. We show that the difference can partially be explained if the correlated volume fluctuations of the solvent are taken into consideration. A special attention is given to the compressibility of proteins. Finally, the present paper is not intended to replace established approaches, but it wants to create awareness that the classical mixing rules refer to ideal gasses assuming uncorrelated volume fluctuations and that a considerable part of the hydration effects could be explained by correlated volume fluctuations.
    Biochimica et Biophysica Acta 09/2008; 1784(11):1546-51. · 4.66 Impact Factor
  • L Smeller, F Meersman, K Heremans
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    ABSTRACT: Pressure unfolding-refolding and the subsequent aggregation of human serum albumin (HSA) was investigated by high-pressure Fourier transform infrared measurements. HSA is completely unfolded at 1 GPa pressure, but the unfolding is not cooperative. Hydrogen-deuterium exchange experiments suggest that a molten globule-like conformation is adopted above 0.4 GPa. An intermediate was formed after decompression, which differs from the native state only slightly in terms of the secondary structure, but this intermediate is more stable against the temperature-induced gel formation than the pressure-untreated native protein. This observation can be explained by assuming that the pressure unfolded-refolded protein is in a misfolded state, which is more stable than the native one.
    European Biophysics Journal 03/2008; 37(7):1127-32. · 2.27 Impact Factor
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    ABSTRACT: High hydrostatic pressure induces conformational changes in proteins ranging from compression of the molecules to loss of native structure. In this tutorial review we describe how the interplay between the volume change and the compressibility leads to pressure-induced unfolding of proteins and dissociation of amyloid fibrils. We also discuss the effect of pressure on protein folding and free energy landscapes. From a molecular viewpoint, pressure effects can be rationalised in terms of packing and hydration of proteins.
    Chemical Society Reviews 11/2006; 35(10):908-17. · 24.89 Impact Factor
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    ABSTRACT: Measurements of residual enzyme activity and FTIR-spectroscopy revealed that mushroom polyphenoloxidase is a thermosensitive enzyme readily inactivated by temperatures exceeding 50°C. The enzyme is, however, very pressure stable. At room temperature no enzyme inactivation occurred at pressures up to 6 kbar. FTIR-spectroscopy revealed that a modification in enzyme conformation occurred below 9 kbar. Kinetic studies showed that pressure and temperature did not always act synergistically with respect to enzyme inactivation. FTIR-spectroscopy showed that pressure- or temperature-induced changes were considerably different. Both temperature and pressure-temperature inactivation were only slightly influenced by pH (in the pH-range 5.5–7.5).
    Journal of Food Science 07/2006; 62(2):261 - 266. · 1.78 Impact Factor
  • Els Loozen, Erik Nies, Karel Heremans, Hugo Berghmans
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    ABSTRACT: In mixtures of PVME and water, the influence of pressure on the LCST miscibility gap is determined covering the whole composition range and pressures from atmospheric pressure up to 900 MPa. The cloud point curve at atmospheric pressure has the characteristic bimodal shape in agreement with literature data. Upon increasing pressure the cloud point curve at the low concentration side decreases with pressure, whereas at the high concentrations the cloud point curve increases with pressure. The overall influence of pressure results in a less pronounced bimodality and ultimately the bimodal shape disappears. In addition to the pressure dependence of the miscibility behavior, the density of mixtures of water and PVME are determined at atmospheric pressure. The experimental excess specific volumes are negative for all measured compositions, but the compositional curvature varies with composition. The curvature of the excess specific volume is positive for the higher concentrations but it is negative in the lower composition range. The density measurements are linked to the pressure dependence of the LCST miscibility behavior using exact thermodynamic relationships. The excess specific volume and miscibility results are shown to be in good agreement. Moreover, it is shown that the Clapeyron equation, which is exact for pure components and also frequently assumed to apply to mixtures, is not valid in the system PVME/water. The system PVME/water is an example where the usual approximation of one-to-one correspondence between curvature and excess volume does not apply. Finally, the molecular origins for the observed excess volume and miscibility behavior are briefly discussed from theoretical and molecular simulation points of view.
    The Journal of Physical Chemistry B 05/2006; 110(15):7793-802. · 3.61 Impact Factor
  • Filip Meersman, László Smeller, Karel Heremans
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    ABSTRACT: The pressure-temperature stability diagram of proteins and the underlying assumptions of the elliptical shape of the diagram are discussed. Possible extensions, such as aggregation and fibril formation, are considered. An important experimental observation is the extreme pressure stability of the mature fibrils. Molecular origins of the diagram in terms of models of the partial molar volume of a protein focus on cavities and hydration. Changes in thermal expansivity, compressibility and heat capacity in terms of fluctuations of the enthalpy and volume change of the unfolding should also focus on these parameters. It is argued that the study of water-soluble polymers might further our understanding of the stability diagram. Whereas the role of water in protein behaviour is unquestioned, the role of cavities is less clear.
    Biochimica et Biophysica Acta 04/2006; 1764(3):346-54. · 4.66 Impact Factor
  • L Smeller, F Meersman, K Heremans
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    ABSTRACT: Refolding of hen egg white lysozyme after pressure unfolding was measured by FTIR spectroscopy. The high-pressure treatment was found to be useful for unfolding/refolding studies because pressure acts against aggregation, and therefore no irreversible aggregation takes place during the pressure treatment. After the release of the pressure, folding intermediate structures were found which were formed during the decompression of the lysozyme. These were aggregation prone when heated, as indicated by their lower stability against aggregation. The intermediates were only formed if the protein was unfolded, subdenaturing pressures could not populate these intermediates. We introduced the notion of a superfunnel to describe the free energy landscape of interacting polypeptide chains. This can explain the propensity of folding intermediates to aggregate. A possible Gibbs-free energy landscape for lysozyme was constructed for the whole pressure-temperature plane.
    Biochimica et Biophysica Acta 04/2006; 1764(3):497-505. · 4.66 Impact Factor
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    H Pfeiffer, G Klose, K Heremans, C Glorieux
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    ABSTRACT: The present paper reports on the phase behaviour of the pseudobinary aqueous mixtures of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC)/pentaethylene glycol monododecyl ether (C12E5) and 1,2-dimyristoyl-sn-glycero-3-phosphocholine monohydrate (DMPC)/C12E5. Both systems exhibit a variety of mesophases, such as lamellar gel, liquid crystalline and micellar phases. The phase diagrams show peritectic and eutectic behaviours. The existence of a compound complex is established. From the phase diagrams, the temperature dependence of the solubilisation parameters is obtained. The phase diagrams, especially with respect to the solubilisation process were qualitatively explained assuming that the packing of the constituents plays a dominating role. Finally, differential scanning calorimetry and ultrasonic velocimetry are compared concerning their potentials to determine characteristics of phase transitions in pseudobinary phospholipid/surfactant mixtures.
    Chemistry and Physics of Lipids 02/2006; 139(1):54-67. · 2.15 Impact Factor
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    ABSTRACT: The changes in hydration of poly(N-isopropylacrylamide) in aqueous solution have been monitored by FTIR spectroscopy as a function of temperature and pressure. Heating the sample above the LCST induces a phase separation that reflects a dehydration of the polymer chain on the molecular level, whereas above the LCSP the phase separation is associated with increased chain hydration. Using these data, we demonstrate the principle of hybrid 2D correlation analysis for the case of two independent data sets obtained under two different physical perturbations. For comparison we also have examined the individual processes by sample–sample (SS) two-dimensional (2D) correlation spectroscopy (COS). SS hybrid 2D correlation analysis enables one to explore the correlation between the concentration dynamics of PNiPA species induced by pressure and temperature, whereas homo SS 2D COS investigates the concentration dynamics of the PNiPA species induced by either one of these physical perturbations. The synchronous SS hybrid-2D correlation spectrum shows a strong negative correlation in the high pressure–temperature region, indicating the pressure- and temperature-induced species are different. Variable–variable (VV) hybrid-2D spectrum is applied to obtain information pertaining to the correlation of conformational variations of PNiPA during the transitions. A critical examination of the method and its applicability is given.
    Journal of Molecular Structure 01/2006; 799:134-140. · 1.40 Impact Factor
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    ABSTRACT: The stability of recombinant Aspergillus aculeatus PME (pectin methylesterase), an enzyme with high beta-helix content, was studied as a function of pressure and temperature. The conformational stability was monitored using FTIR (Fourier transform IR) spectroscopy whereas the functional enzyme stability was monitored by inactivation studies. Protein unfolding followed by amorphous aggregation, which makes the process irreversible, was observed at temperatures above 50 degrees C. This could be correlated to the irreversible enzyme inactivation observed at that temperature. Hydrostatic pressure greater than 1 GPa was necessary to induce changes in the enzyme's secondary structure. No enzyme inactivation was observed at up to 700 MPa. Pressure increased PME stability towards thermal denaturation. At 200 MPa, temperatures above 60 degrees C were necessary to cause significant PME unfolding and loss of activity. These results may be relevant for an understanding of the extreme stability of amyloid fibrils for which beta-helices have been proposed as a structural element.
    Biochemical Journal 01/2006; 392(Pt 3):565-71. · 4.65 Impact Factor

Publication Stats

2k Citations
245.88 Total Impact Points

Institutions

  • 1989–2013
    • University of Leuven
      • • Department of Chemistry
      • • Department of Metallurgy and Materials Engineering
      • • Centre for Food and Microbial Technology
      Louvain, Flanders, Belgium
  • 1999–2008
    • Semmelweis University
      • Department of Biophysics and Radiation Biology
      Budapest, Budapest fovaros, Hungary
    • Academia Sinica
      T’ai-pei, Taipei, Taiwan
  • 2005–2006
    • University of Cambridge
      • Department of Chemistry
      Cambridge, ENG, United Kingdom
  • 2002
    • Unité Inserm U1077
      Caen, Lower Normandy, France
    • INSA
      Альтамира, Tamaulipas, Mexico
  • 2001
    • French National Centre for Scientific Research
      Lutetia Parisorum, Île-de-France, France
  • 1996
    • French Institute of Health and Medical Research
      Lutetia Parisorum, Île-de-France, France
  • 1995
    • Moscow State Forest University
      Mytishi, Moskovskaya, Russia
  • 1988
    • National Research Council Canada
      Ottawa, Ontario, Canada