Assessing Squarates as Amine-Reactive Probes.

Probes that covalently label protein targets facilitate the identification of ligand-binding sites. Lysine residues are prevalent in the proteome, making them attractive substrates for covalent probes. However, identifying electrophiles that undergo amine-specific, regioselective reactions with binding site lysine residues is challenging. Squarates can engage in two sequential conjugate addition-elimination reactions with amines. Nitrogen donation reduces the second reaction rate, making the mono squaramide a mild electrophile. We postulated that this mild electrophilicity would demand a longer residence time near the amine, affording higher selectivity for binding site lysines. Therefore, we compared the kinetics of squarate and monosquaramide amine substitution to alternative amine bioconjugation handles. The data revealed that N-hydroxy succinimidyl esters react 4 orders of magnitude faster, consistent with their labeling promiscuity. Squarate reactivity can be tuned by a substitution pattern. Electron-withdrawing groups on the vinylogous ester or amide increase reaction rates. Dithionosquarates react more rapidly than squarates, while vinylogous thioester analogs, dithiosquarates, react more slowly. We assessed squarate selectively using the UDP-sugar processing enzyme GlfT2 from Mycobacterium tuberculosis, which possesses 21 surface-exposed lysines. The reaction predominately modified one lysine proximal to a binding site to afford covalent inhibition. These findings demonstrate the selectivity of squaric esters and squaramides, which is a critical feature for affinity-based chemoproteomic probes.

Promoting health in type 2 diabetes: nurse-physician collaboration in primary care.

The purpose of this study is to examine effects of a nurse-physician collaborative approach to care of patients with type 2 diabetes and to determine possible effect sizes for use in computing sample sizes for a larger study. Forty patients from a family practice clinic with type 2 diabetes were randomly assigned to control or experimental groups. The control group received standard care, whereas the experimental group received standard care plus home visits from a nurse, as well as consultation with an exercise specialist and/or nutritionist. Follow-up continued for 3 months. Clinical end points included standard measures of diabetes activity as well as quality-of-life indicators. Focus group interviews were used to explore patients' responses to the program. Although findings were not statistically significant, a trend toward small to moderate positive effect sizes was found in glycosylated hemoglobin and blood pressure. Quality of life measures also showed a trend toward small to moderate, but nonsignificant, improvements in physical functioning, bodily pain, vitality, social and global functioning, energy, impact of diabetes, and health distress. Focus group interviews indicated a very positive response from patients, who expressed feelings of empowerment. In this study, patients treated with nurse-physician collaboration demonstrated small, but nonsignificant, improvements in blood chemistry after only 3 months. Physical and social functioning, energy, and bodily pain also showed a small improvement. Changes in awareness of effects of diabetes on health and an expressed sense of self-efficacy suggest that effects could be sustainable over the longer term.