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Polydiacetylene Monolayers: Model Systems for Enzymatic Surface Modification

Published online by Cambridge University Press:  21 February 2011

Troy E. Wilson  and
Mark D. Bednarski
Troy E. Wilson
Affiliation:
Department of Chemistry, University of California at Berkeley and the Center for Advanced Materials, Lawrence Berkeley Laboratory, Berkeley, CA 94720
Mark D. Bednarski
Affiliation:
Department of Chemistry, University of California at Berkeley and the Center for Advanced Materials, Lawrence Berkeley Laboratory, Berkeley, CA 94720
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Extract

We are exploring the requirements for enzyme-catalyzed reactions on small molecules tethered to the surfaces of organic monolayers. Despite considerable effort toward understanding enzymatic processes in solution1, the chemistry of enzymes at interfaces has not been studied. Increasingly sophisticated methods of surface modification, including self-assembly2 and photolithographic3 techniques, raise intriguing prospects for enzymatic surface chemistry. This paper describes our initial investigations of the proteolysis of a dipeptide substrate covalently tethered to the surface of a polydiacetylene film using the enzyme, subtilisin BPN'.

Type
Research Article
Information
MRS Online Proceedings Library (OPL) , Volume 237: Symposium Ca – Interface Dynamics and Growth , 1991 , 317
Copyright
Copyright © Materials Research Society 1992

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References

1. Fersht, A., Enzyme Structure and Mechanism, 2nd ed. (W. H. Freeman Publishers, New York, 1985).Google Scholar
2. For reviews see: (a) Bain, C.D. and Whitesides, G.M., Adv. Mater. 4, 110 (1989).Google Scholar
(b) Swalen, J.D., Allara, D.L., Andrade, J.D., Chandross, E.A., Garoff, S., Israelachvili, J., McCarthy, T.J., Murray, R., Pease, R.F., Rabolt, J.F., Wynne, K.J., Yu, H., Langmuir 3, 932 (1987).Google Scholar
(c) Ulman, A., J. Mat. Educ. 11, 205 (1989).Google Scholar
3. Fodor, S.P.A., Read, J.L., Pirrung, M.C., Stryer, L., Lu, A.-T., Solas, D., Science 251, 767 (1991).Google Scholar
4. Wilson, T.E. and Bednarski, M.D., Langmuir (in press).Google Scholar
5. Nagy, J.O. and Bednarski, M.D., manuscript in preparation.Google Scholar
6. Kraut, J., Ann. Rev. Biochem. 46, 331 (1977).CrossRefGoogle Scholar
7. The synthesis and analytical data for compound 1 will be published elsewhere.Google Scholar
8. The glass microscope slides were made hydrophobic by pre-treatment with octadecylsilane following the method of Maoz, R. and Sagiv, J., Thin Solid Films 132, 135 (1985).Google Scholar
9. Subtilisin BPN' (P-4789, lot 39F0458) and succinyl-L-alanyl-L-alanyl-L-prolyl-L-phenylalaninep-nitroanilide (sAAPFpna) were purchased from Sigma Chemical and used as received. The enzyme was assayed for activity prior to use and all quantities of enzyme refer to active enzyme content.Google Scholar
10. The N-terminal benzoyl amide bond has been shown not to be a substrate for the subtilisin BPN' protease. For a more detailed discussion, see Bender, M.L. and Kezdy, F.J., Ann. Rev. Biochem. 34, 49 (1965).CrossRefGoogle Scholar
11. XPS spectra were obtained using a Phi 5300 spectrometer (Perkin-Elmer Instruments) with MgKcc radiation of 1253.6 eV, quartz monochromator, concentric hemispherical analyzer operating in fixed analyzer transmission mode and multichannel detector. The base pressure in the chamber was approximately 1 × 10-8 Torr. Survey spectra were recorded with a 45° takeoff angle, 180-eV pass energy, 0.500 eV/step, 20-mm2 spot and 400-W electron beam power with an acquisition time of 10 min. High-resolution spectra were recorded with an 18-eV pass energy, 1 mm-spot and 400-W power. Signals were referenced to the Ag3d5/2 peak at 367.9 eV. For an excellent treatment of XPS, see:Google Scholar
Practical Surface Analysis by Auger and X-ray Photoelectron Spectroscopy, 2nd ed., edited by Briggs, D. and Seah, M.P. (Wiley and Sons, Chichester, U.K., 1990), Vol. 1.Google Scholar
12. N-Tosyl-L-phenylalanine chloromethyl ketone (TPCK) was obtained from Boehringer Mannheim Biochemicals (Indianapolis, IN) and was used following the procedure of Schoellmann, G. and Shaw, E., Biochemistry 2, 252 (1963).Google Scholar
13. The X-ray Photoelectron Spectrometer (XPS) measures the chemical composition of the topmost 50–100 Å of the target surface. The presence of “buried” F5-Bz-Phe-OH moieties in the polydiacetylene film would be detected. For further information, see 11.Google Scholar