Defining Resonance Raman Spectral Responses to Substrate Binding by Cytochrome P450 from \u3cem\u3ePseudomonas putida\u3c/em\u3e

Resonance Raman spectra are reported for substrate-free and camphor-bound cytochrome P450cam and its isotopically labeled analogues that have been reconstituted with protoheme derivatives that bear -CD3 groups at the 1, 3, 5, and 8-positions (d12-protoheme) or deuterated methine carbons (d4-protoheme). In agreement with previous studies of this and similar enzymes, substrate binding induces changes in the high frequency and low frequency spectral regions, with the most dramatic effect in the low frequency region being activation of a new mode near 367 cm−1. This substrate-activated mode had been previously assigned as a second “propionate bending” mode (Chen et al., Biochemistry, 2004, 43, 1798–1808), arising in addition to the single propionate bending mode observed for the substrate-free form at 380 cm−1. In this work, this newly activated mode is observed to shift by 8 cm−1 to lower frequency in the d12-protoheme reconstituted enzyme (i.e., the same shift as that observed for the higher frequency “propionate bending” mode) and is therefore consistent with the suggested assignment. However, the newly acquired data for the d4-protoheme substituted analogue also support an earlier alternate suggestion (Deng et al., Biochemistry, 1999, 38, 13699–13706) that substrate binding activates several heme out-of-plane modes, one of which (γ6) is accidentally degenerate with the 367 cm−1 propionate bending mode. Finally, the study of the enzyme reconstituted with the protoheme-d4, which shifts the macrocycle ν10 mode, has now allowed a definitive identification of the vinyl CC stretching modes

The Integrated Spectra of M32 and of 47 Tucanae: A Comparative Study in the Mid-Ultraviolet With IUE

Low-resolution mid-UV spectra of M32 and 47 Tuc have been extracted from the IUE archival database, along with spectra of 41 F and G dwarfs with well-determined atmospheric parameters and integrated spectra of 24 Galactic globular clusters. We have used five mid-UV spectral indices defined by Fanelli et al. to constrain the stellar content of M32 and 47 Tuc and to make a comparative study between the two stellar systems. In the case of 47 Tuc, the bulk of the mid-UV light is shown to come from the main-sequence turnoff stars, with much smaller (but significant) contributions coming from red horizontal-branch stars, red giants, and A stars (presumably, blue stragglers). In contrast, M32 is shown to have no significant contribution from a red horizontal-branch population, has a more metal-rich main-sequence turnoff, and has a significantly larger hot star contribution than is inferred to be present in 47 Tuc. These inferences are consistent with conclusions obtained from integrated light studies of M32 and 47 Tuc in the blue

Quantification of uf and pf Resins in MDF Fiber with an X-Ray Fluorescence Spectrometer

This article describes methods to quantify urea-formaldehyde (UF) resin and phenol-formaldehyde (PF) resin contents in medium density fiberboard (MDF) using a wavelength dispersive X-ray fluorescence spectrometer (WDXRF). The methods are based on the principle that a specific metallic element shows its characteristic X-ray fluorescence spectrum, the intensity of which is correlated quantitatively to its concentration. In the case of UF-blended MDF fiber, 2.4% copper sulfate pentahydrate CuSO4.5H2O (based on resin solids) was premixed with the resin as a labeling agent. Quantification of copper ion was performed using XRF. Based on calibrations with laboratory-prepared standard fiber samples of known UF resin and copper quantities, the results of XRF measurements were converted to resin loading rates. In the case of PF-blended fiber, the PF resin contents in the MDF fiber samples were successfully quantified by measuring the existing sodium ion Na+ in the resol resin with XRF. Linear calibration curves between fluorescence intensity of copper or sodium and resin content were established respectively for UF and PF resins. Test results show that the methods were precise and reliable

Resonance Raman Detection of the Hydroperoxo Intermediate in the Cytochrome P450 Enzymatic Cycle

The resonance Raman spectra of the hydroperoxo complex of camphor-bound CYP101 have been obtained by cryoradiolytic reduction of the oxygenated ferrous form that had been rapidly frozen in water/glycerol frozen solution; EPR spectroscopy was employed to confirm the identity of the trapped intermediate. The ν(O−O) mode, appearing at 799 cm-1, is observed for the first time in a peroxo-heme adduct. It is assigned unambiguously by employing isotopomeric mixtures of oxygen gas containing 50% 16O18O, confirming the presence of an intact O−O fragment. The ν(Fe−O) mode is observed at 559 cm-1 (H2O). Furthermore, both modes shift down by 3 cm-1, documenting the formulation as a hydroperoxo complex, in agreement with EPR data

Effects of Hot-Pressing Parameters and Wax Content on the Properties of Fiberboard Made from Paper Mill Sludge

Primary sludge combined with 20% secondary sludge was used for the manufacture of fiberboard. A factorial design was carried out to determine the effects of panel density, pressing temperature and time, and wax level on the panel properties of fiberboard. Two levels were employed for each of the four variables, and the panel dimensional stability and mechanical properties were analyzed using Design-Expert software. The statistical analysis indicated that internal bonding (IB) was significantly affected by panel density, pressing temperature, and their interaction. Pressing time and wax level were not directly related to IB. Similarly, modulus of rupture (MOR) was dependent strongly on panel density, pressing temperature, and their interaction, but was not affected by pressing time and wax level. The effect of panel density on modulus of elasticity (MOE) was as strong as on MOR, but the effect of pressing temperature was weaker on MOE than on MOR. MOE was also related to pressing time, but not to wax level. Thickness swelling (TS) was not affected by panel density, but it was significantly dependent on pressing temperature and time. Unexpectedly, wax level did not have significant impact on TS

The Anodic Dissolution Of Cadmium

The anodic dissolution of cadmium has been studied in aqueous solutions containing Cl−, Br−, I−, Ac−, SO4=, and NO3− ions. The normal valence (+2) was found in all solutions with the exception of NO3−. The apparent valence (calculated) of cadmium ions in nitrate solutions varied from 1.2 to 2.0 and was found to be a function of NO3− concentration, current density, and temperature. An anodic dissolution mechanism has been proposed involving local corrosion and disintegration of the anode which is consistent with the experimental results. © 1967, The Electrochemical Society, Inc. All rights reserved

Early changes in pro-inflammatory cytokine levels in neonates with encephalopathy are associated with remote epilepsy.

BackgroundNeonatal seizures are associated with adverse neurologic sequelae including epilepsy in childhood. Here we aim to determine whether levels of cytokines in neonates with brain injury are associated with acute symptomatic seizures or remote epilepsy.MethodsThis is a cohort study of term newborns with encephalopathy at UCSF between 10/1993 and 1/2000 who had dried blood spots. Maternal, perinatal/postnatal, neuroimaging, and epilepsy variables were abstracted by chart review. Logistic regression was used to compare levels of cytokines with acute seizures and the development of epilepsy.ResultsIn a cohort of 26 newborns with neonatal encephalopathy at risk for hypoxic ischemic encephalopathy with blood spots for analysis, diffuse alterations in both pro- and anti-inflammatory cytokine levels were observed between those with (11/28, 39%) and without acute symptomatic seizures. Seventeen of the 26 (63%) patients had >2 years of follow-up and 4/17 (24%) developed epilepsy. Higher levels of pro-inflammatory cytokines IL-6 and TNF-α within the IL-1β pathway were significantly associated with epilepsy.ConclusionsElevations in pro-inflammatory cytokines in the IL-1β pathway were associated with later onset of epilepsy. Larger cohort studies are needed to confirm the predictive value of these circulating biomarkers