In Rhizopus, membrane Repotrectinib ligand-binding assays suggest the presence of a progesterone receptor but that has not led to the identification of the specific receptor [27–30]. In this work we identified a homologue of the PAQR family as an interacting protein of the CBL0137 solubility dmso S. schenckii G protein alpha

subunit, SSG2, using the yeast two-hybrid analysis. Using a yeast-based assay we determined that progesterone was the ligand of this S. schenckii PAQR (SsPAQR1). This assay was used because it is specific for PAQRs and was intended for the study of these receptors without the intervention of other possible progesterone binding proteins. The receptor was expressed in S. cerevisiae that has no other known progesterone receptor. We also report the effects of this agonist on the growth of the fungus buy SIS3 from conidia and on the intracellular cyclic 3′, 5′ adenosine monophosphate (cAMP) levels in S. schenckii yeast cells at various time intervals following exposure to the hormone. Results Yeast two-hybrid screening A yeast two-hybrid assay was done using the complete coding sequence of SSG-2 as bait and a S. schenckii yeast cells cDNA library. In this screening,

a 483 bp insert from a blue colony growing in quadruple drop out (QDO) medium (SD/-Ade/-His/-Leu/-Trp/X-α-gal) was sequenced and found to encode the last 38 amino acid of the C-terminal residues of a protein homologous to Izh3 from S. cerevisiae (GenBank no. NP_013123.1). Sequencing of the SsPAQR1 gene Figure1 shows the cDNA and derived amino acid sequence of sspaqr1 gene obtained using 5′ RACE. This figure shows a 1981 bp cDNA with an ORF of 1542 bp encoding a 514 amino acid protein with a calculated molecular

weight of 57.8 kDa. The GenBank accession numbers for the cDNA and derived amino acid sequence, respectively are: EU439945.1 and ACA43006.1. The PANTHER Classification System identified this protein as a member of the PAQR family (PTHR20855:SF10) (residues 149-512) with an extremely significant E value of 3.8 e-158[31]. Figure 1 cDNA and derived amino acid sequences of the sspaqr1 gene. Figure1A shows the hemolysin III motif identified using the NCBI Conserved Domain Database. The hemolysin III motif Venetoclax research buy that is present in all PAQRs, extends from amino acid 270 to 495. Figure1B shows the cDNA and derived amino acid sequence of the sspaqr1 gene. Non-coding regions are given in lower case letters, coding regions and amino acids are given in upper case letters. Motif A, B and C are shaded yellow, blue-green, and green, respectively. Motif C includes part of the original sequence isolated in the yeast two-hybrid assay. The original sequence isolated using the yeast two-hybrid assay is underlined. Figure1 also shows the characteristic residues that identify the members of the Class II PAQR family of receptors. The Class II PAQR family (progesterone receptors) is characterized by the presence of 7 transmembrane domains, and three highly conserved amino acid motifs [13].