The Amidase Domain of Lipoamidase Specifically Inactivates Lipoylated Proteins In Vivo

BACKGROUND:In the 1950s, Reed and coworkers discovered an enzyme activity in Streptococcus faecalis (Enterococcus faecalis) extracts that inactivated the Escherichia. coli and E. faecalis pyruvate dehydrogenase complexes through cleavage of the lipoamide bond. The enzyme that caused this lipoamidase activity remained unidentified until Jiang and Cronan discovered the gene encoding lipoamidase (Lpa) through the screening of an expression library. Subsequent cloning and characterization of the recombinant enzyme revealed that lipoamidase is an 80 kDa protein composed of an amidase domain containing a classic Ser-Ser-Lys catalytic triad and a carboxy-terminal domain of unknown function. Here, we show that the amidase domain can be used as an in vivo probe which specifically inactivates lipoylated enzymes. METHODOLOGY/PRINCIPAL FINDINGS:We evaluated whether Lpa could function as an inducible probe of alpha-ketoacid dehydrogenase inactivation using E. coli as a model system. Lpa expression resulted in cleavage of lipoic acid from the three lipoylated proteins expressed in E. coli, but did not result in cleavage of biotin from the sole biotinylated protein, the biotin carboxyl carrier protein. When expressed in lipoylation deficient E. coli, Lpa is not toxic, indicating that Lpa does not interfere with any other critical metabolic pathways. When truncated to the amidase domain, Lpa retained lipoamidase activity without acquiring biotinidase activity, indicating that the carboxy-terminal domain is not essential for substrate recognition or function. Substitution of any of the three catalytic triad amino acids with alanine produced inactive Lpa proteins. CONCLUSIONS/SIGNIFICANCE:The enzyme lipoamidase is active against a broad range of lipoylated proteins in vivo, but does not affect the growth of lipoylation deficient E. coli. Lpa can be truncated to 60% of its original size with only a partial loss of activity, resulting in a smaller probe that can be used to study the effects of alpha-ketoacid dehydrogenase inactivation in vivo

A Pilot Study of Self-Reported Physical Activity and Eating Habits in Turkish Cancer Patients Under Chemotherapy

As in all individuals, improving the quality of life, balanced nutrition and physical activity habits must be acquired in cancer patients. The purpose of this study was to determine eating habits and physical activity of cancer patients receiving chemotherapy. Sixty-six patients were completed the questionnaire included sociodemographic data, type of cancer, anthropometric measurements (size and body weight), dietary and physical activity habits. Body mass index for each patient was calculated. Data were analyzed using Statistical Package for Social Science software. Patients were ranged from underweight to obese according to their body mass index: 6.1% of patients were classified as underweight. Almost half (48.5%, n = 32) reported to be regularly physical active, and 46.9% (n = 15) thereof reported 30min brisk walking. More vegetables consumption was the most popular answer with 62.1% (n = 41), whereas vegetables/fruit or vegetables/legume consumption was 22.7% (n = 15). Gender differences in food choice and preferring the taste of food were not seen as statistically significant. In this article, patients with different types of cancer reported their eating habits and physical activity. Disease-related and worse prognostic factors were found. An institutional program should be offered to cancer patients for consulting about nutrition and physical activity