We demonstrate that the steady state reaction of lysine 5,6-aminomutase with substrate analogue 4-thia-l-lysine generates a radical intermediate, which accumulates in the enzyme to an electron paramagnetic resonance (EPR) detectable level. EPR line width narrowing of approximately 1 mT due to [4'-(2)H] labeling of the pyridoxal-5'-phosphate (PLP), an isotropic hyperfine coupling of 40 MHz for the proton at C4' of PLP derived from (2)H electron nuclear double resonance (ENDOR) measurement, and spin density delocalization onto the (31)P of PLP realized from observations of the (31)P ENDOR signal provide unequivocal identification of the radical as a substrate-PLP-based species. X- and Q-band EPR spectra fittings demonstrate that this radical is spin coupled with the low spin Co(2+) in cob (II) alamin and the distance between the two species is about 10 A. These results provide direct evidence for the active site motion upon substrate binding, bringing the adenosylcobalamin to the proximity of substrate-PLP for subsequent H-atom abstraction and for the notion that lysine 5,6-aminomutase functions by a radical mechanism. Observation of (2)H-ENDOR signal also provides a reliable hyperfine coupling constant for future comparison with quantum-mechanical-based calculations to gain further insight into the molecular structure of this steady state radical intermediate.

We performed spectroscopic and functional characterization on cyanobacterium Synechocystis PCC6803 with mutations of charged residues of the cytoplasmic side of cytochrome (Cyt) b559 in photosystem II (PSII). All of the mutant cells grew photoautotrophically and assembled stable PSII. However, R7Eα, R17Eα and R17Lβ mutant cells grew significantly slower and were more susceptible to photoinhibition than wild-type cells. The adverse effects of the arginine mutations on the activity and the stability of PSII were in the following order (R17Lβ>R7Eα>R17Eα and R17Aα). All these arginine mutants exhibited normal period-four oscillation in oxygen yield. Thermoluminescence characteristics indicated a slight decrease in the stability of the S3QB(-)/S2QB(-) charge pairs in the R7Eα and R17Lβ mutant cells. R7Eα and R17Lβ PSII core complexes contained predominantly the low potential form of Cyt b559. EPR results indicated the displacement of one of the two axial ligands to the heme of Cyt b559 in R7Eα and R17Lβ mutant reaction centers. Our results demonstrate that the electrostatic interactions between these arginine residues and the heme propionates of Cyt b559 are important to the structure and redox properties of Cyt b559. In addition, the blue light-induced nonphotochemical quenching was significantly attenuated and its recovery was accelerated in the R7Lα and R17Lβ mutant cells. Furthermore, ultra performance liquid chromatography-mass spectrometry results showed that the PQ pool was more reduced in the R7Eα and R17Lβ mutant cells than wild-type cells in the dark. Our data support a functional role of Cyt b559 in protection of PSII under photoinhibition conditions in vivo.

The paper proposes a utility function and game theory (UGT) based network selection scheme to maximize accommodated number of calls, minimize handoff occurrence frequency, and fulfill quality of service (QoS) requirements in heterogeneous wireless networks. When a new call or a handoff call arrives, UGT would calculate the utility value and preference value for each candidate network based on the QoS satisfaction of the call request and the cooperative game computation. Finally, the network, which has the maximum result of linearly combined utility value and preference value, would be selected as the most suitable network for the call request. Simulation results show that UGT has good QoS satisfaction. It also allows system to accommodate more calls than iterative TOPSIS scheme. In addition, UGT can reduce the handoff occurrence frequency by about 30% as compared to iterative TOPSIS.

Adenosylcobalamin- and pyridoxal-5'-phosphate-dependent lysine 5,6-aminomutase utilizes free radical intermediates to mediate 1,2-amino group rearrangement, during which an elusive high-energy aziridincarbinyl radical is proposed to be central in the mechanism of action. Understanding how the enzyme participates in stabilizing any of the radical intermediates is fundamentally significant. Y263F mutation abolished the enzymatic activity. With isotope-edited EPR methods, the roles of the Tyr263α residue in the putative active site are revealed. The Tyr263α residue stabilizes a radical intermediate, which most likely is the aziridincarbinyl radical, either by acting as a spin-relay device or serving as an anchor for the pyridine ring of pyridoxal-5'-phosphate through aromatic π-stacking interactions during spin transfer. The Tyr263α residue also protects the radical intermediate from interception by molecular oxygen. This study supports the proposed reaction mechanism, including the aziridincarbinyl radical, which has eluded detection for more than two decades.