The potential of perturbed angular correlation of gamma rays as a tool for dynamic studies of peptides/proteins

Bauer, Rogert; Atke, A.; Danielsen, Eva; Marcussen, Jan; Olsen, Carl Erik; Jf, Rehfeld; Særmark, T.; Schneider, Dietmar; Vilhardt, Hans; Zeppezauer, Michael

doi:10.1016/0883-2889(91)90003-j

Cited by 14 publications

(9 citation statements)

References 23 publications

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“…All water was distilled twice, and adjustment of pH was done by using either NaOH or sulfuric acid in order to avoid Cl™. All glassware was washed in aqua regia, and all water was passed through a diethylenetriaminepentaacetate-coupled gel (height = 3 cm, diameter = 1 cm) (Bauer et al, 1991b) in order to lower the contamination of metals to less than 0.1 µ .…”

Section: Methodsmentioning

confidence: 99%

Cd-substituted horse liver alcohol dehydrogenase: catalytic site metal coordination geometry and protein conformation

Hemmingsen¹,

Bauer²,

Mj³

et al. 1995

Biochemistry

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The coordination geometry of the catalytic site in Cd-substituted horse liver alcohol dehydrogenase (LADH) has been investigated as a function of pH using the method of perturbed angular correlation of gamma-rays (PAC). LADH in solution fully loaded with cadmium, including radioactive 111mCd in the catalytic site [Cd2(111mCd)Cd2LADH], was studied over the pH range 7.9-11.5. Analysis of the PAC spectra showed the ionization of a group with pKa of 11. This pKa value is about 2 pH units higher than that of native zinc-containing LADH. A pKa of 9.6 was found for the binary complex of Cd2(111mCd)Cd2LADH with NAD+. This value is also about 2 pH units higher than that of the binary complex of native zinc-containing enzyme and NAD+. No pH dependency was detected for the binary complex of Cd2(111mCd)Cd2LADH with NADH within the pH range measured (pH 8.3-11.5). Assuming that metal-coordinated water is the ionizing group [Kvassman, J., & Pettersson, G. (1979) Eur. J. Biochem. 100, 115-123], we conclude that the larger ionic radius of Cd(II) relative to Zn(II) in the catalytic site causes the elevated pKa values of metal-bound water. Interpretation of nuclear quadrupole interaction (NQI) parameters derived from PAC spectra is based on the use of the angular overlap model, using the coordinates for the catalytic zinc site from the 1.8 A resolution crystal structure of the ternary complex between LADH, NADH, and dimethyl sulfoxide as a model.(ABSTRACT TRUNCATED AT 250 WORDS)

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Section: Methodsmentioning

confidence: 99%

Cd-substituted horse liver alcohol dehydrogenase: catalytic site metal coordination geometry and protein conformation

Hemmingsen¹,

Bauer²,

Mj³

et al. 1995

Biochemistry

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“…The spectral parameters for all the complexes of LADH with coenzyme and/or inhibitors investigated by PAC (Andersson et al, 1982;Bauer et al, 1991 a;Hemmingsen et al, 1995; this work) fall into one of two groups: a low o,, region (120-230 Mrad/s) or a high o, region (270-350 Mrad/s); 9 is relatively constant in all experiments. From the results in this work it has become clear that there is a connection between the presence of a charged solvent ligand and spectral parameters in the low w, region.…”

Section: Fractionmentioning

confidence: 99%

The Protein Conformation of Cd‐Substituted Horse Liver Alcohol Dehydrogenase and its Metal‐Site Coordination Geometry in Binary and Ternary Inhibitor Complexes

Hemmingsen

Bauer

Bjerrum

et al. 1996

European Journal of Biochemistry

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The coordination geometry of the metal at the active site in Cd-substituted horse liver alcohol dehydrogenase (LADH) has been investigated for the binary complexes of LADH with imidazole, isobutyramide, decanoic acid and C1-, and for the ternary complexes of LADH with NADH and imidazole, NADH and isobutyramide, NAD' and decanoic acid and NAD' and Cl-, by using the method of perturbed angular correlation of y-rays (PAC). The spectral results are consistent with a flexible structure around the metal for the binary complexes with inhibitors. For ternary complexes, however, a rigid structure is observed. An exception is the ternary complex between LADH, NADH and imidazole, in which the metal site is still flexible. Comparing with available structures determined by X-ray crystallography, we found a correlation between open structures and flexible metal sites, and between closed structures and rigid metal sites. This indicates that the PAC technique can be applied to distinguish the two conformations in solution.The spectral parameters, oo and v, of the experiments, except for the complexes with imidazole, fall into two groups: one with low oo and one with high o, (7 is relatively constant in all experiments). In this work it is clarified that the low o,, values are connected with the presence of a negatively charged solvent ligand. Using an angular-overlap approach to interpret the results, the low cc),, values are found to be compatible with a coordination geometry where the S-Cd-S (Cys174 and Cys46 coordinate to the metal) angle is about 110" as suggested in [Hemmingsen, L., Bauer, R., Danielsen, E., Bjerrum, M. J., Zeppezauer, M., Adolph, H. W., Formicka, G. & Cedergren-Zeppezauer, E. (1995) Biochemistry 34, 7145571531, whereas high coo values are compatible with an S-Cd-S angle of 130". The presence of a negatively charged metal ligand, therefore, might trigger the movement of the sulfur of Cys174. As it is believed that alcohols coordinate to the metal as alcoholate ions this could be important for catalysis.Keywords: perturbed angular correlation ; protein conformation ; metal-site structure; function ; flexibility.Horse liver alcohol dehydrogenase (LADH), a dimeric Zndependent enzyme, catalyses the reversible oxidation of primary and secondary alcohols using NAD' as coenzyme. A TheorellChance model is assumed for the catalytic process (Kvassman

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“…This is mainly due to the low concentrations of the radiometals and chelators. However, the common methods of determination are thin layer chromatography and high-performance liquid chromatography, in addition to further animal studies. , PAC is a useful technique for these purposes, as is presented in a number of works. − However, it has not become a routine technique, mainly due to uncertainty concerning after-effects, the need for a convenient parameter, and a relatively small choice of the radionuclides suitable for PAC. The aim of the present work is to provide an example of the application of PAC and its suitability for the study of a chelator–radionuclide system.…”

Section: Introductionmentioning

confidence: 99%

“…The γ–γ perturbed angular correlation technique is based on the angular correlation between two γ-rays, which is perturbed by the hyperfine interaction of the nuclear electric quadrupole moment ( Q ) with the electric field gradient (EFG) or the nuclear magnetic moment with an extranuclear magnetic field. PAC gives information serving as a “fingerprint” of the local structure. − As the PAC can be used to measure changes in the local environment of the nucleus, this technique can be useful for more detailed investigation of the radionuclides attached to a ligand …”

Section: Introductionmentioning

confidence: 99%

“…20−22 As the PAC can be used to measure changes in the local environment of the nucleus, this technique can be useful for more detailed investigation of the radionuclides attached to a ligand. 5 It is known that electron capture (EC) decay creates a hole in one of the inner shells (K, L), which moves toward the outer shells due to Auger-electron and X-ray emissions. 23 The Auger cascade becomes a reason for the daughter to be left in a highly ionized state, which may cause changes at the probe site, which in turn would cause changes in the static and dynamic characteristics of the electric field.…”

Section: ■ Introductionmentioning

confidence: 99%

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Perturbed Angular Correlation as a Tool to Study Precursors for Radiopharmaceuticals

et al. 2020

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One of the key components of radiopharmaceuticals for targeting imaging and therapy is a stable bifunctional chelating system to attach radionuclides to selective delivery systems. After-effects of radioactive decay can cause the release of a radioactive isotope from its chelation agent. Perturbed angular correlation (PAC) of γ-rays has become a unique technique to study the behavior of complexes formed between a chelating agent and radionuclide in vivo (in real time) over a relevant range of concentrations (10 −12 M). In the present work, four radionuclides, 111 In, 111m Cd, and 152, 154 Eu, were investigated with diethylenetriaminepentaacetic acid (DTPA) at different pH values to determine the stability constants of the complexes as well as the effects of post-decay processes, which play a major role in determining the suitability of these complexes for application as radiopharmaceuticals (e.g., in vivo generators). The study provides a convenient parameter for the characterization of radionuclide−chelator systems using the PAC method. PAC is proven to be a suitable tool to study novel chelators and radiopharmaceutical precursors attached to radiometals.

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The potential of perturbed angular correlation of gamma rays as a tool for dynamic studies of peptides/proteins

Cited by 14 publications

References 23 publications

Cd-substituted horse liver alcohol dehydrogenase: catalytic site metal coordination geometry and protein conformation

Cd-substituted horse liver alcohol dehydrogenase: catalytic site metal coordination geometry and protein conformation

The Protein Conformation of Cd‐Substituted Horse Liver Alcohol Dehydrogenase and its Metal‐Site Coordination Geometry in Binary and Ternary Inhibitor Complexes

Perturbed Angular Correlation as a Tool to Study Precursors for Radiopharmaceuticals

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