No abstract

Hepatic‐type fatty‐acid‐binding protein (hFABP) from the cytosol of bovine liver is a 14.4‐kDa neutral protein with a blocked N‐terminus and a disulfide system located on the surface of the protein. It binds two molecules of fatty acid in one binding site, apparent dissociation constants of the oleic acid/hFABP complex are 0.24 μM and 2.15 μM. Computer analysis of circular dichroic spectra predicts that hFABP contains about 12%α‐helix, 45%β‐structure, 15%β‐turn and 27% unordered structure. Ellipticities indicative of secondary structure are not affected by fatty acid binding. Cationic amino acid residues of hFABP (1 His, 15 Lys, 2 Arg) were screened for ionic fatty acid/protein interactions. His was excluded, as 1H‐NMR analysis of His‐C2 and His‐C4 protons indicated that binding of oleic acid shifts the pK of His from 6.9 to 7.1 only in hFABP with the disulfide system in the oxidized state; acylation of His with diethylpyrocarbonate does not affect the binding of the fatty acid. Acetylation of Lys reduces binding marginally, whereas modification of Arg with phenylglyoxal lowers the binding activity by 65%. From 1H‐NMR investigations, conformational changes within the protein, due to a sort of disaggregation of hFABP upon fatty acid binding, were derived. Most of the proton resonances sharpen up with ligand binding, and some of the methyl resonances shift positions, possibly because they are directly involved in the fatty acid/protein interaction.

Our data indicate a more rapid clot lysis at low UK and rt-PA concentrations in newborns despite significantly reduced plasminogen levels. The results of the plasmin generation experiments suggest a diminished effect of plasmin inhibitors towards fetal plasmin, which raises an explanation for the concentration-related differences in the clot lysis assay. The experience with thrombolytic agents in newborns is limited. Most dosage regimens for thrombolytic therapy in children or adults consist of an initial bolus infusion, followed by low-dose continuous treatment. Based on the results of our clot lysis experiments, we think that especially the continuous infusion of plasminogen activators after bolus administration should not be enhanced in newborns compared to older children or adults.

The fluorescence decay of chlorophyll in spinach thylakoids was measured as a function of the degree of closure of Photosystem II reaction centers, which was set for the flowed sample by varying either the preillumination by actinic light or the exposure of the sample to the exciting pulsed laser light. Three exponential kinetic components originating in Photosystem II were fitted to the decays; a fourth component arising from Photosystem I was determined to be negligible at the emission wavelength of 685 nm at which the fluorescence decays were measured. Both the lifetimes and the amplitudes of the components vary with reaction center closure. A fast (170-330 ps) component reflects the trapping kinetics of open Photosystem II reaction centers capable of reducing the plastoquinone pool; its amplitude decreases gradually with trap closure, which is incompatible with the concept of photosynthetic unit connectivity where excitation energy which encounters a closed trap can find a different, possibly open one. For a connected system, the amplitude of the fast fluorescence component is expected to remain constant. The slow component (1.7-3.0 ns) is virtually absent when the reaction centers are open, and its growth is attributable to the appearance of closed centers. The middle component (0.4-1.7 ns) with approximately constant amplitude may originate from centers that are not functionally linked to the plastoquinone pool. To explain the continuous increase in the lifetimes of all three components upon reaction center closure, we propose that the transmembrane electric field generated by photosynthetic turnover modulates the trapping kinetics in Photosystem II and thereby affects the excited state lifetime in the antenna in the trap-limited case.