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Bio-orthogonal copper (I)-catalyzed azide–alkyne cycloaddition (CuAAC) has been widely used to modify azide- or alkyne-bearing monosaccharides on metabolic glyco-engineered mammalian cells. Here, we present a systematic study to elucidate the design space for the cytotoxic effects of the copper catalyst on NIH 3T3 fibroblasts and on HEK 293-F cells. Monitoring membrane integrity by flow cytometry and RT-PCR analysis with apoptotic and anti-apoptotic markers elucidated the general feasibility of CuAAC, with exposure time of the CuAAC reaction mixture having the major influence on biocompatibility. A high labeling efficiency of HEK 293-F cells with a fluorescent alkyne dye was rapidly achieved by CuAAC in comparison to copper free strain-promoted azide–alkyne cycloaddition (SPAAC). The study details effective and biocompatible conditions for CuAAC-based modification of glyco-engineered cells in comparison to its copper free alternative.

Metabolic glyco-engineering is a widely used technique to modify glyco-structures on living cells. We report a design space for efficient, rapid, and cell-compatible surface modification by comparing bio-orthogonal copper-catalyzed azide–alkyne cycloaddition with its copper-free alternative.

CDC via Flickr, CC by ND 2.0

E. coli bacteria

If infected with E. coli, some people have worse symptoms than others. New research suggests that varying gene expression might be the reason why.

You probably know that your DNA contains the blueprints for every protein your body needs, plus instructions on how to regulate them. But your DNA could also strongly influence your immune system, making you more or less susceptible to bacterial infections, according to a new study led by researchers from Duke University. The study was published this week in the Journal of Infectious Diseases.

Scientists have already had some inklings of the relationship between genes and infectious diseases—for example, people with the genetic mutation for cystic fibrosis don’t usually get typhoid, which is caused by bacteria. Bacteria and viruses, in turn, can also affect your genes.

For this study, the researchers wanted to see if genes affected a person’s likelihood of contracting a common bacterial infection. The researchers infected 30 participants with E. coli bacteria, a common cause of diarrhea. For the next eight days, the researchers watched for symptoms—six participants basically showed no symptoms, while another six were debilitated by the infection—and then drew their blood.

The researchers were checking the blood for gene expression. While every cell might have thousands of genes, only a select few are activated at any given time, turned on throughout the process of development or by external factors like smoking and diet.

Bacteria, it seems, can also modify gene expression—when the researchers compared the gene expression of participants with severe symptoms and those with few symptoms, they found significant differences in the expression of 29 genes related to immune function. It seems that certain genes were turned on when the bacteria were present, making the participants more immune. They anticipate that those variations could help predict which patients will react strongly to an E. coli infection.

What's not clear, though, is if the participants with few symptoms had mutations in those particular genes, or if those genes reacted more strongly to the presence of the bacteria. To answer that question and to further confirm their findings, the researchers hope to perform similar experiments with other types of bacteria and viruses, paying special attention to those 29 genes they suspect play a role in disease resistance. If they’re right, it’s possible that someday, treatments for infectious diseases might rely more on epigenetics, activating infection-resistant genes so that patients suffer less.