Small GTP-binding proteins in the nuclei of human placenta

Mitochondria and crude nuclei containing fractions from human Placenta have been shown to contain proteins which bind [alpha P-32]-GTP. Prior to this study the number of GTP-binding proteins in placental nuclei and their nucleotide specificity was not known. Also unknown was the identity of any of the GTP-binding proteins in mitochondria of human placenta. Nuclei and mitochondria were purified from human placental extracts by sedimentation. Proteins were separated by electrophoresis and transferred to nitrocellulose membranes. Overlay blot with [alpha P-32]-GTP identified two nuclei proteins with approximate molecular weights of 24 and 27 kDa. Binding of [alpha P-32]-GTP to the 27 and 24 kDa proteins was significantly displaced by guanine nucleotides but not by adenine, thymine or cytosine nucleotides or deoxy (d) GTP. Western blot with a specific antibody to Ran identified a band at 27 kDa in nuclei and in mitochondrial fractions. These data indicate that both nuclei and mitochondria contain 24 and 27 kDa GTP-binding proteins. The CTP-binding proteins in nuclei display binding specificity fur guanine nucleotides 27 kDa GTP-binding and the hydroxylated carbon 2 on the ribose ring of GTP appears essential for binding. It will be important in future studies to determine the functions of these small GTP-binding proteins in the development and physiology of the placenta. (C) 2001 Wiley-Liss, Inc.