Effect of Interaction Between Methotrexate and Dihydrofolate Reductase on DNA Synthesis in L1210 Cells In Vitro

L1210 leukaemia cells were preloaded with [3H methotrexate] (MTX) to saturate high-affinity intracellular sites, and were then incubated with [3H]MTX to determine the steady-state intracellular MTX concentrations at extracellular concentrations ranging from 10 μM to zero. In addition, incubations to generate incomplete saturation of high-affinity intracellular MTX-binding sites were also carried out. Following determination of the total intracellular MTX, cells were pulsed with deoxyuridine (UdR) and its incorporation into DNA examined to assess the role of exchangeable and bound intracellular MTX on DNA synthesis. Further, cell pellets were disrupted and dihydrofolate reductase (DHFR) activity determined under each experimental condition. Extracellular MTX concentrations in excess of 1 μM depressed UdR incorporation to <2% of control, but incorporation rapidly recovered to 62% of control at the point of MTX-DHFR equivalence, and exceeded control values when all high-affinity sites were not saturated. DHFR activity was undetectable at extracellular MTX concentrations >0.50 μM, and never exceeded 6.09% of control at the “equivalence point” where all high-affinity sites were saturated. When less than 10% of potential inhibitor sites were occupied, enzyme activity increased rapidly, but never reached control. However, 5% of the DHFR activity was sufficient to permit UdR incorporation to continue at 50% of control levels, and UdR incorporation returned to control levels at 20% of the DHFR activity. The relationship between cellular MTX content and DNA synthesis or DHFR activity is sigmoid, suggesting a reversible interaction between enzyme and inhibitor. This lends support to the notion that “free” intracellular MTX is necessary for a maximal antitumour effect, and may explain its role in “high-dose” MTX therapy in man

Learning by Doing: Performance Assessment of Information Literacy across the First-Year Curriculum

In the 2011-2012 school year, the instruction librarians at Augustana College, Illinois, changed their assessment approach in the college’s required first-year sequence to focus on higher-level information literacy concepts. The librarians replaced a quantitative assessment instrument with performance assessments, which they integrated into their first-year library sessions. Although the sequence is taught by many faculty with diverse assignments, these new assessments could be applied organically across sections yet provide generalizable results. This case study describes that assessment project and its initial findings, analyzes the project’s implications, and suggests how other college libraries might adopt similar qualitative assessments

A new restriction endonuclease from the anaerobic bacterium, Desulfovibrio desulfuricans, Norway.

The purification and characterization of a new restriction endonuclease, Dde 1, from a sulfate-reducing, anaerobic bacterium, Desulfovibrio desulfuricans, Norway, is reported. The enzyme recognizes the sequence (see formula index) and cleaves at the position indicated by the arrows. The enzyme preparation obtained is suitable for restriction mapping an ligation