Bradykinin and its gly sup 6 analogue are substrates of cyclophilin: A fluorine-19 magnetization transfer study

Journal Article · Thu Nov 01 04:00:00 EST 1990 · Biochemistry; (United States)

DOI:https://doi.org/10.1021/bi00497a002· OSTI ID:5557213

London, R E; Davis, D G ^[1]; Vavrek, R J; Stewart, J M ^[2]; Handschumacher, R E ^[3]

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1. NIEHS, NC (USA)
2. Univ. of Colorado Health Sciences Center, Denver, CO (USA)
3. Yale Univ., New Haven, CT (USA)

Fluorine-19 magnetization transfer experiments have been used to determine the rates of cis/trans isomerization about the X-Pro{sup 7} peptide bond in (p-fluoro-Phe{sup 8})bradykinin and its Gly{sup 6} analogue. The measurements were carried out both prior to and after the addition of cyclophilin, which has recently been shown to have peptidyl-proline cis/trans isomerase activity and is the apparent target enzyme of the immunosuppressive agent cyclosporin A. Magnetization transfer measurements over the temperature range 40-75 {degree}C in the absence of enzyme give activation energies of 22.8 and 23.0 kcal/mol for (p-fluoro-Phe{sup 8})bradykinin and its Gly{sup 6} analogue, respectively. The values for the uncatalyzed cis {r arrow} trans rate constant, k{sub c}, are determined by extrapolation to be 4.8 {times} 10{sup {minus}2} and 2.1 {times} 10{sup {minus}2} s{sup {minus}1} for the two peptides at 25 {degree}C. The enzyme-catalyzed enhancement of the cis/trans interconversion rate was proportional to added cyclophilin concentration and was strongly sequence specific, with bradykinin a much better substrate than (Gly{sup 6})bradykinin. At a peptide concentration of 2.2 mM, the catalytic activity expressed as k{sub c} per micromolar cyclophilin was determined to be 1.2 s{sup {minus}1}/{mu}M for (p-fluoro-Phe{sup 8})bradykinin and 0.13 s{sup {minus}1}/{mu}M for the Gly{sup 6}analogue. The increased cis {r arrow} trans interconversion rates were strongly inhibited by cyclosporin A and the 6-(methylalanine) derivative, which bind to cyclophilin, but not by the 1-(tetrahydrofurfuryl) derivative of cyclosporin that binds weakly.

• OSTI ID: 5557213
• Journal Information: Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 29:45; ISSN 0006-2960; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACIDS
AMINOBUTYRIC ACID
ARRHENIUS EQUATION
AUTONOMIC NERVOUS SYSTEM AGENTS
BIOCHEMICAL REACTION KINETICS
CARBOXYLIC ACIDS
DRUGS
ENZYME ACTIVITY
ENZYME INHIBITORS
ENZYMES
EQUATIONS
FLUORINE 19
FLUORINE ISOTOPES
GLYCINE
HYDROXY ACIDS
ISOMERASES
ISOTOPES
KINETICS
KININS
LIGHT NUCLEI
MAGNETIC RESONANCE
NEUROREGULATORS
NUCLEAR MAGNETIC RESONANCE
NUCLEI
ODD-EVEN NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
PEPTIDES
POLYPEPTIDES
PROTEIN STRUCTURE
PROTEINS
REACTION KINETICS
RELAXATION
RESONANCE
SERINE
SPIN-LATTICE RELAXATION
STABLE ISOTOPES