Adapting recombinant bacterial alkaline phosphatase for nucleotide exchange of small GTPases.

The small GTPase Rat sarcoma virus proteins (RAS) are key regulators of cell growth and involved in 20-30% of cancers. RAS switches between its active state and inactive state via exchange of GTP (active) and GDP (inactive). Therefore, to study active protein, it needs to undergo nucleotide exchange to a non-hydrolysable GTP analog. Calf intestine alkaline phosphatase bound to agarose beads (CIP-agarose) is regularly used in a nucleotide exchange protocol to replace GDP with a non-hydrolysable analog. Due to pandemic supply problems and product shortages, we found the need for an alternative to this commercially available product. Here we describe how we generated a bacterial alkaline phosphatase (BAP) with an affinity tag bound to an agarose bead. This BAP completely exchanges the nucleotide in our samples, thereby demonstrating an alternative to the commercially available product using generally available laboratory equipment.

Chylous Cardiac Tamponade with Chylothoraces Secondary to Hodgkin's Lymphoma: Octreotide in Conjuncture with Standard of Care Dietary Fat Restriction.

Chylous effusions are a well-known complication from a variety of etiologies including trauma, malignancies, and anatomic defects, with the most common location being in the pleural space. A pericardial chylous effusion (chylopericardium) is uncommon, and a chylopericardium with concomitant bilateral chylous pleural effusions (chylothoraces) has only been reported in less than a handful of case reports. Our patient presented with bilateral chylothoraces and a chylopericardium with tamponade physiology secondary to Hodgkin's Lymphoma. In this article, we discuss our treatment of this patient with the somatostatin analogue octreotide, as well as the standard of care dietary fat restriction, in order to control these effusions until the patient's chemotherapy took effect.