BIOIiVORGANIC
CHEMISTR
Y $21-32
21
(1975)
X-Ray Photadectron Spectroscopy of Some SeleniumContaining Amino Acids HEINZ
RUPP and ULRICH
WESER
Physiologisch-chemisches Institut der UniversirZt D-74 Tiibingen, Hoppe-Seyler Str. I, Germany
Tiibingen
ABSTRACT X-ray photoelectron spectra of some inorganic selenium compounds, Se-methionine, Secystine, Se -ur= and selenodicysteine were recorded and compared with the XPS data obtained from the respective sulphur containing compounds. The oxidation state of selenium could be monitored by the observed chemical shifts of the Se(3pl@, Se(3p3/2) and Se(3d3/2,5,2) levels. Though having a formal oxidation state near zero, the binding energy of the core electrons of Se in Se-methionine, Se-cystine and selenodicysteine was shifted by 0.4, 0.7 and 0.4 eV, respectively. This phenomenon was attributed to the rather distinct polarization of Se. The reversible oxidation of Secystine using HZ@, and NaBH, could be successfully demonstrated by this XPS-technique. KEY WORDS Selenium, X-ray photoelectron urea, Selenodicysteine.
spectroscopy,
Se-cystine, Se-methionine, Se-
INTRODUCTION Almost 20 years have passed since selenium was elucidated as an essential and biochemically active trace element [ l]_ Apart from its wide distribution in biological tissues [2], selenium proved strongly involved in various enzymic processes where it was found in stoichiometric amounts [ 3-6) _ Furthermore, low molecular weight selenium compounds can activate a number of enzymes [7-S I _ Due to its redox sensitive character, it was presumed that selenium can be regarded to represent the vital portion of the active site in the above selenium enzymes. Although this explanation seems rather attractive no final decision can be made at the moment_ An approach in this direction was the assignment of the oxidation state of selenium in biological material avoiding its chemical destruction prior to the respective measurements_ For this purpose X-ray photoelectron spectroscopy turned out to be most suitable since the chemical shift of the binding energy of the core electrons was expected to be rather distinct in the differently oxidized selenium compounds_ The energy values of
., 0 American Elsevier Publishing Company, Inc., 1975
XPS OF SELENO AMINO ACIDS
31
state was obtained_ The quality of the XPS spectrum of selenodicysteine was extraordinarily high. The complete symmetry and the small full width at half maximum indicate the absence of selenium in another oxidation state. Regarding the oxidation state of selenium iu selenodicysteiue au interesting conclusion‘ could be made. Favoring the Pauling electronegativity scale 1221, it is proposed that sulphur is slightly more electronegative compared to selenium. This codd canse a shift of the binding energies of the selenium core electrons to somewhat higher values. In fact, the energy values of the selenium in selenodicysteine is shifted by +0.4 eV compared to elemental selenium_ iu a higher oxidation
CONCLUSION
The X-ray photoelectron spectroscopic data obtained from some organoselenium compounds [2 t ] and, as in the present case, using seleno amino acids are very promising. The marked chemical shift of the different oxidized and reduced selenium species makes XI% spectroscopy to a powerful tool for studying selenoenzymes. However, a snfficiently high sensitivity of the employed instrumentation is required. This condition was already fulfilled in earlier XPS measurements using ferredoxin, erythrocuprein (superoxide dismutase) and metallothionein [ 13-18]_ In these meta? proteins both the presence and oxidation state of sulphur, iron, copper, zinc, cadmium and mercury were successfully demonstrated.
HR. is a recipient of a preabctoral grant of the Stiftung Villigst. This study was aided by DFG-grants No. We 40119 and 401/11 awarded to U-it! T/ze skillful assistance of Mrs. K. Rupp is greatly appreciated
REFERENCES 1. K. Schwarz and C. M. FoItz,J.
Amer. Chem. Sot
79,3292-3293
(1957)_
2. W. J. Rhead, G. A. Evans, and G_ N. Schrauzer,Bioinorg. Chem. 3,217-223
(1974). 3. L. Flohe, W. A. Giinzler, and H. H. &hock, FEBS Letters 32,132-134 (1973). 4_ W_ G.. Hoekstra, in Proceedings of the Second International Symposium on ZTace Element Metabolism in Animals. Univ. Park Press, Baltimore, 1974, pp. 61-77. 5. P. D. Whanger, N. D. Pedersen, and P. H. Weswig, Biochem. Biophys. Res. Commun. 53, 1031-1035 (1973). 6. D. C. Turner and T. C. Stadtman,Ar&. Biochem. Biophys. 154,366-381 (1973). 7. R. C. Dicksonand A. L. Tappel, Arch. Biochem. Biophys. 131,100-110 (1969). 8. A. T. Diplock and J. A. Lucy, FEBS Letters 29,205-210 (1973). 9. H. Rupp, Biochemical Master Thesis, Tiibingen, 1973. 10. H. E. Ganther,Eiochemistry 7,2898-2905 (1968). Ii. K_ Siegbahn, C. Nordling, A. Fahlman, R. Nordberg, K. Hamrin, J_ Hedman, G. Johansson, T. Bergmark, S. E. Karlsson, L. Lindgren, and B. Lindberg, ESCA -Atomic Molecular and Solid State Structures Studied by Means of Electron Spectroscopy, Almqvist and WikseiIs; Uppsala, 1967.
32
HEINZ RUPP AND ULRICH
WESER
12, C. K. J$rgensen, C?zimia25,213-222 (1971). 13. L. N. Kramer and M. P. IUein, in Electron Spectroscopy (D. A_ Shirley, Ed..), North-HolIand, Amsterdam, 1972. pp. 733-751. 14. G. Jung, M. Ottuad, W. Bohenkamp, and U. Weser, FEBSLerrers 25,346-348 (1972). 15. 6. Jung, M. Ottnad, Vf_ Bohnenkamp, W_ Bremser, and U. Weszr, Biochim. Biophys. Acto 295,?7-86 (1973)_ 16. U. Weser, H. Rupp, F_ Donay, F. Lixmemarm, W. Voelter, W. Voetsch, Eur_ J. Biochem. 39,127-140 (1973).
and G_ Jung,
17. H. Rupp and U. Weser, FEBS Letter? 44,293-297 (1974)18. G. Sokoloxvski, W_ Pilz, md U_ Weser, FEBS Letters, 48, 222-225 (1974). 19. C_ K_ Jbrgensen and H. Berthou, Photo-Electron Spectra induced by X-rays of Above 600 Non-i%etaItic Compounds Conrating 77 Elements, Munksgaard, Kobenham, 1972. 20_ W. E. Swartz, K_ J. Wynne, and D. M. Hercules, Anal. Chem. 43,1884-l 887 (1971).
21. G_ MaImsten, I_ Thorin, S. Hiigberg, J. E. Bergmark, S. E. Karlsson, and E. Rebane, Pizysira Scripta 3,96-100 (1971). 22 L. PauIing, The IVatwe of the chemical Bond, Cornell Univ. Press, Ithaca, 1960_ 23. C. B. Knobier, Y. Okaya, and R. Pepinsky, 2. Kn‘stallogr 111,385397 (1959). 24_ D_ M_ Hercules,AnaL Cbem_ 42 (1) 20A (1970). 25. K. A_ Caldwell and A. L. Tappel, Biochemistry 3,1643-1647 (1964). Received IS October IV -4; revised I6 December I974
CHEMISTR
Y $21-32
21
(1975)
X-Ray Photadectron Spectroscopy of Some SeleniumContaining Amino Acids HEINZ
RUPP and ULRICH
WESER
Physiologisch-chemisches Institut der UniversirZt D-74 Tiibingen, Hoppe-Seyler Str. I, Germany
Tiibingen
ABSTRACT X-ray photoelectron spectra of some inorganic selenium compounds, Se-methionine, Secystine, Se -ur= and selenodicysteine were recorded and compared with the XPS data obtained from the respective sulphur containing compounds. The oxidation state of selenium could be monitored by the observed chemical shifts of the Se(3pl@, Se(3p3/2) and Se(3d3/2,5,2) levels. Though having a formal oxidation state near zero, the binding energy of the core electrons of Se in Se-methionine, Se-cystine and selenodicysteine was shifted by 0.4, 0.7 and 0.4 eV, respectively. This phenomenon was attributed to the rather distinct polarization of Se. The reversible oxidation of Secystine using HZ@, and NaBH, could be successfully demonstrated by this XPS-technique. KEY WORDS Selenium, X-ray photoelectron urea, Selenodicysteine.
spectroscopy,
Se-cystine, Se-methionine, Se-
INTRODUCTION Almost 20 years have passed since selenium was elucidated as an essential and biochemically active trace element [ l]_ Apart from its wide distribution in biological tissues [2], selenium proved strongly involved in various enzymic processes where it was found in stoichiometric amounts [ 3-6) _ Furthermore, low molecular weight selenium compounds can activate a number of enzymes [7-S I _ Due to its redox sensitive character, it was presumed that selenium can be regarded to represent the vital portion of the active site in the above selenium enzymes. Although this explanation seems rather attractive no final decision can be made at the moment_ An approach in this direction was the assignment of the oxidation state of selenium in biological material avoiding its chemical destruction prior to the respective measurements_ For this purpose X-ray photoelectron spectroscopy turned out to be most suitable since the chemical shift of the binding energy of the core electrons was expected to be rather distinct in the differently oxidized selenium compounds_ The energy values of
., 0 American Elsevier Publishing Company, Inc., 1975
XPS OF SELENO AMINO ACIDS
31
state was obtained_ The quality of the XPS spectrum of selenodicysteine was extraordinarily high. The complete symmetry and the small full width at half maximum indicate the absence of selenium in another oxidation state. Regarding the oxidation state of selenium iu selenodicysteiue au interesting conclusion‘ could be made. Favoring the Pauling electronegativity scale 1221, it is proposed that sulphur is slightly more electronegative compared to selenium. This codd canse a shift of the binding energies of the selenium core electrons to somewhat higher values. In fact, the energy values of the selenium in selenodicysteine is shifted by +0.4 eV compared to elemental selenium_ iu a higher oxidation
CONCLUSION
The X-ray photoelectron spectroscopic data obtained from some organoselenium compounds [2 t ] and, as in the present case, using seleno amino acids are very promising. The marked chemical shift of the different oxidized and reduced selenium species makes XI% spectroscopy to a powerful tool for studying selenoenzymes. However, a snfficiently high sensitivity of the employed instrumentation is required. This condition was already fulfilled in earlier XPS measurements using ferredoxin, erythrocuprein (superoxide dismutase) and metallothionein [ 13-18]_ In these meta? proteins both the presence and oxidation state of sulphur, iron, copper, zinc, cadmium and mercury were successfully demonstrated.
HR. is a recipient of a preabctoral grant of the Stiftung Villigst. This study was aided by DFG-grants No. We 40119 and 401/11 awarded to U-it! T/ze skillful assistance of Mrs. K. Rupp is greatly appreciated
REFERENCES 1. K. Schwarz and C. M. FoItz,J.
Amer. Chem. Sot
79,3292-3293
(1957)_
2. W. J. Rhead, G. A. Evans, and G_ N. Schrauzer,Bioinorg. Chem. 3,217-223
(1974). 3. L. Flohe, W. A. Giinzler, and H. H. &hock, FEBS Letters 32,132-134 (1973). 4_ W_ G.. Hoekstra, in Proceedings of the Second International Symposium on ZTace Element Metabolism in Animals. Univ. Park Press, Baltimore, 1974, pp. 61-77. 5. P. D. Whanger, N. D. Pedersen, and P. H. Weswig, Biochem. Biophys. Res. Commun. 53, 1031-1035 (1973). 6. D. C. Turner and T. C. Stadtman,Ar&. Biochem. Biophys. 154,366-381 (1973). 7. R. C. Dicksonand A. L. Tappel, Arch. Biochem. Biophys. 131,100-110 (1969). 8. A. T. Diplock and J. A. Lucy, FEBS Letters 29,205-210 (1973). 9. H. Rupp, Biochemical Master Thesis, Tiibingen, 1973. 10. H. E. Ganther,Eiochemistry 7,2898-2905 (1968). Ii. K_ Siegbahn, C. Nordling, A. Fahlman, R. Nordberg, K. Hamrin, J_ Hedman, G. Johansson, T. Bergmark, S. E. Karlsson, L. Lindgren, and B. Lindberg, ESCA -Atomic Molecular and Solid State Structures Studied by Means of Electron Spectroscopy, Almqvist and WikseiIs; Uppsala, 1967.
32
HEINZ RUPP AND ULRICH
WESER
12, C. K. J$rgensen, C?zimia25,213-222 (1971). 13. L. N. Kramer and M. P. IUein, in Electron Spectroscopy (D. A_ Shirley, Ed..), North-HolIand, Amsterdam, 1972. pp. 733-751. 14. G. Jung, M. Ottuad, W. Bohenkamp, and U. Weser, FEBSLerrers 25,346-348 (1972). 15. 6. Jung, M. Ottnad, Vf_ Bohnenkamp, W_ Bremser, and U. Weszr, Biochim. Biophys. Acto 295,?7-86 (1973)_ 16. U. Weser, H. Rupp, F_ Donay, F. Lixmemarm, W. Voelter, W. Voetsch, Eur_ J. Biochem. 39,127-140 (1973).
and G_ Jung,
17. H. Rupp and U. Weser, FEBS Letter? 44,293-297 (1974)18. G. Sokoloxvski, W_ Pilz, md U_ Weser, FEBS Letters, 48, 222-225 (1974). 19. C_ K_ Jbrgensen and H. Berthou, Photo-Electron Spectra induced by X-rays of Above 600 Non-i%etaItic Compounds Conrating 77 Elements, Munksgaard, Kobenham, 1972. 20_ W. E. Swartz, K_ J. Wynne, and D. M. Hercules, Anal. Chem. 43,1884-l 887 (1971).
21. G_ MaImsten, I_ Thorin, S. Hiigberg, J. E. Bergmark, S. E. Karlsson, and E. Rebane, Pizysira Scripta 3,96-100 (1971). 22 L. PauIing, The IVatwe of the chemical Bond, Cornell Univ. Press, Ithaca, 1960_ 23. C. B. Knobier, Y. Okaya, and R. Pepinsky, 2. Kn‘stallogr 111,385397 (1959). 24_ D_ M_ Hercules,AnaL Cbem_ 42 (1) 20A (1970). 25. K. A_ Caldwell and A. L. Tappel, Biochemistry 3,1643-1647 (1964). Received IS October IV -4; revised I6 December I974
