Isolation and characterization of a cDNA clone encoding a member of the Com44/Cim44 envelope components of the chloroplast protein import apparatus

Abstract

Many of the proteins in the chloroplast envelope play an important role in facilitating the biochemical and transport processes of the compartment. For the transport of proteins into the chloroplast, we have recently identified at least three different envelope proteins (Com44/Cim44, Com70, and Cim97) in close physical proximity to a partially translocated chimeric precursor protein (Wu, C, Seibert, F. S., and Ko, K. (1994) J. Biol. Chem. 269, 32264- 32271). In this study we report the characterization of a cDNA clone encoding a member of the Com44/Cim44 envelope proteins. The combined data from nucleotide sequencing, and RNA and protein blot analyses indicate the existence of multiple forms of the 44-kDa envelope protein. Depending on the plant species examined, immunologically-related protein bands with molecular masses of 42 to 46 kDa were observed. Organelle subfractionation, protease treatment, and immunomicroscopic studies together provide an indication that the immunologically-related proteins may be present in both the outer and inner envelope membranes. Co-migration of the product synthesized from the cDNA insert with a 44-kDa immunoreactive band of the chloroplast envelope, and the in vitro import results, together suggest that the in vitro synthesized 44-kDa protein is targeted to the envelope membrane without any further processing.