J. Korean Soc. Appl. Biol. Chem. 53(6), 852-856 (2010) Short Communication Crystal Structure of a Putative Histidine-containing Phosphotransfer Protein from Oryza sativa Euiyoung Bae 1,2 , Eduard Bitto 1,3 , Craig A. Bingman 1 , Simon T. Allard 1 , Gary E. Wesenberg 1 , Russell L. Wrobel 1 , Brian G. Fox 1 , and George N. Phillips, Jr. 1 * 1 Department of Biochemistry and Center for Eukaryotic Structural Genomics, University of Wisconsin-Madison, Madison, Wisconsin, 53706, USA 2 Department of Agricultural Biotechnology and Center for Agricultural Biomaterials, Seoul National University, Seoul 151-921, Republic of Korea 3 Department of Chemistry and Biochemistry, Georgian Court University, Lakewood, New Jersey, 18701, USA Received August 31, 2010; Accepted September 7, 2010 Os09g0567400 codes for a hypothetical protein from Oryza sativa that is annotated as the “Histidine-containing phosphotransfer (Hpt) protein”. Hpt domain is a protein module with a histidine residue mediating phosphotransfer reaction in the histidine-aspartate phosphorelay system. We report here the crystal structure and analysis of Os09g0567400. Key words: histidine-containing phosphotransfer protein, histidine-aspartate phosphorelay system, Os09g0567400 Os09g0567400 codes for a hypothetical protein from Oryza sativa that is annotated in the UniProt knowledgebase [Jain et al., 2009] as the “Histidine-containing phosphotransfer (Hpt) protein”. Its molecular weight is 16.8 kDa, and it comprises 149 amino acid residues. BLAST searches revealed homologues of Os09g0567400 in many plants including Zea mays and Arabidopsis thaliana [Altschul et al., 1997]. The homologues and Os09g0567400 were identified as members of the Pfam family, Hpt (PF01627) [Bateman et al., 2004], which is named after the Hpt domain, a protein module with a histidine residue mediating phosphotransfer reaction in the histidine-aspartate phosphorelay system [Kato et al., 1997; Porter et al., 2003]. The histidine-aspartate phosphorelay is a signal transduction pathway originally found in bacteria [Stock et al., 2000], and also identified in fungi and plants [Saito, 2001]. In plants, the histidine-aspartate phosphorelay systems are comprised of three protein components; a sensory histidine kinase, a Hpt protein, and a response regulator. First, the sensory histidine kinase catalyzes a phosphorylation reaction of its own histidine residue and subsequently transfers the phosphoryl group to an aspartate residue of its internal receiver domain. It is then moved to a histidine residue of the Hpt protein, and finally delivered to an aspartate residue of the response regulator. This system has been extensively studied in A. thaliana, with the homologous Hpt proteins known as AHP1-6, corresponding to gene loci At3g21510, At3g29350, At5g39340, At3g11360, At1g03430, and At1g80100, respectively. These proteins participate in the cytokinin signal transduction pathway, where histidine protein kinases (AHKs) act as receptors and the AHPs (Htps) transmit the signal to response regulators in the nucleus (ARRs) [Muller and Sheen, 2007]. The annotation of Os09g0567400 as a rice Hpt protein is based on amino acid sequence comparison, and its biological function or three-dimensional structure has not been previously characterized experimentally. In eukaryotes, crystal structures of the Hpt proteins have been determined for Saccharomyces cerevisiae and Z. mays [Song et al., 1999; Sugawara et al ., 2005]. Homologues were originally chosen for study from A. thaliana, but these proteins failed at various steps in our pipeline [Thao et al., 2004; Jeon et al., 2005; Sreenath et al., 2005]. To demonstrate the usefulness of the rice genome for structural genomics activities on higher plants and to add examples of this important class of proteins, the crystal structure and analysis of Os09g0567400, UniGene code Os.4702, UniProt code Q6VAK4 are reported here. *Corresponding author Phone: +1-608-263-6142; Fax: +1-608-262-3453 E-mail: phillips@biochem.wisc.edu doi:10.3839/jksabc.2010.130