Activity in monomers of human cytomegalovirus protease.

Herpesvirus proteases require dimerization for activity, although crystallographic data indicate that each monomeric subunit possesses a well-separated and complete active site. This suggests that dimerization stabilizes the monomeric protease subunits in an active conformation. Chemical cross-linking with disuccinimidyl glutarate was used to capture human cytomegalovirus protease in its various conformations. The cross-linked protease retained activity under mildly chaotropic conditions (0.25 to 0.75 M urea) in contrast to non-cross-linked protease which lost activity. Identification of active protease species by incorporation of radioactive diisopropylfluorophosphate showed that in addition to cross-linked dimers, cross-linked protease monomers were responsible for a significant fraction of the total protease activity. These results are consistent with the hypothesis that herpesvirus protease activation occurs by stabilization of an active conformer in the dimer.     

Bordetella pertussis Serotypes in the United States

To determine the Bordetella pertussis serotypes currently causing whooping cough in the United States, recently isolated cultures from different geographic areas were studied. Specific adsorbed antisera were prepared in our laboratory and used in both tube and slide agglutination tests. Among the 177 cultures isolated during 1966 and 1967 in seven states and one Canadian province, the predominant serotype was 1.3 (or 1.3.6), represented by 85% of the cultures. Fifteen cultures were serotype 1.2.3.4.6. The 376 cultures isolated in the Grand Rapids area during the last 30 years showed a changing pattern from serotype 1.2.3 during the early years to the currently prevailing 1.3 (or 1.3.6). During the 2-year study period, seven cultures of B. parapertussis were isolated in Michigan, and seven were received from other states. One culture of B. bronchiseptica, recovered from a child in Boston, was sent to us.     

Spectroscopic studies of stellacyanin derivatives

Two covalently modified derivatives of the apoprotein of the blue copper protein stellacyanin have been prepared. In one case, a dansyl group was linked to the cysteine at the copper binding site of apostellacyanin; in the other, a nitrophenol moiety has been attached to this same cysteine. Fluorescence yields and emission maxima of the dansylated protein and pK determinations of the nitrophenol group linked to the protein suggest that the solvent microenvironment at the copper binding site of apostellacyanin is quite similar to bulk water.     

Knockout of cGAS and STING Rescues Virus Infection of Plasmid DNA-Transfected Cells

It is well known that plasmid DNA transfection, prior to virus infection, negatively affects infection efficiency. Here, we show that cytosolic plasmid DNA activates the cGAS/STING signaling pathway, which ultimately leads to the induction of an antiviral state of the cells. Using a transient one-plasmid clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 system, we generated cGAS/STING-knockout cells and show that these cells can be infected after plasmid DNA transfection as efficiently as nontransfected cells.     

Closing the carbon balance for fermentation by Clostridium thermocellum (ATCC 27405)

Our lab and most others have not been able to close a carbon balance for fermentation by the thermophilic, cellulolytic anaerobe, Clostridium thermocellum. We undertook a detailed accounting of product formation in C. thermocellum ATCC 27405. Elemental analysis revealed that for both cellulose (Avicel) and cellobiose, >92% of the substrate carbon utilized could be accounted for in the pellet, supernatant and off-gas when including sampling. However, 11.1% of the original substrate carbon was found in the liquid phase and not in the form of commonly-measured fermentation products - ethanol, acetate, lactate, and formate. Further detailed analysis revealed all the products to be <720 da and have not usually been associated with C. thermocellum fermentation, including malate, pyruvate, uracil, soluble glucans, and extracellular free amino acids. By accounting for these products, 92.9% and 93.2% of the final product carbon was identified during growth on cellobiose and Avicel, respectively.     

The performance of fungal xylan-degrading enzyme preparations in elemental chlorine-free bleaching for Eucalyptus pulp

Cellulase-free xylan-degrading enzyme preparations from Acrophialophora nainiana, Humicola grisea var. thermoidea and two Trichoderma harzianum strains were used as bleaching agents for Eucalyptus kraft pulp, prior to a chlorine dioxide and alkaline bleaching sequence. In comparison to the control sequence (performed without xylanase pretreatment), the sequence incorporating enzyme treatment was more effective. Removal of residual lignin was indicated by a reduction in kappa number. Overall, enzyme preparations from T. harzianum were marginally more effective in reducing pulp viscosity and chlorine chemical consumption and improving the brightness of the kraft pulp. However, the highest reduction in pulp viscosity was mediated by the xylanase preparation from A. nainiana. Xylanase pretreatment compares very favorably with that of chemical pulping. Journal of Industrial Microbiology & Biotechnology (2002) 28, 204–206 DOI: 10.1038/sj/jim/7000227 Received 27 April 2001/ Accepted in revised form 03 November 2001