The psaA and psaB genes, which encode the P700 chlorophyll a apoproteins of the Photosystem I complex, have been cloned from the unicellular, transformable cyanobacterium Synechococcus sp. PCC 7002. The nucleotide sequence of these genes and of their flanking sequences have been determined by the chain termination method. As found in the chloroplast genomes of higher plants, the psaA gene lies 5' to the psaB gene; however, the cyanobacterial genes are separated by a greater distance (173 vs. 25-26 bp). The psaA gene is predicted to encode a polypeptide of 739 amino acid residues (81.7 kDa), and the psaB gene is predicted to encode a polypeptide of 733 residues (81.4 kDa). The cyanobacterial psa gene products are 76% to 81% identical to their higher plant homologues; moreover, because of conservative amino acid replacements, the cyanobacterial sequences are more than 95% homologous to those determined for higher plants. These results provide the basis for a genetic analysis of Photosystem I, and are discussed in relationship to structural and functional aspects of the Photosystem I complexes of both cyanobacteria and higher plants.

No abstract

Eleven structural analogues of human basic fibroblast growth factor (bFGF) have been prepared by site-directed mutagenesis of a synthetic bFGF gene to examine the effect of amino acid substitutions in the three putative heparin-binding domains on FGF's biological activity. After expression in Escherichia coli, the mutant proteins were purified to homogeneity by use of heparin-Sepharose chromatography and analyzed for their ability to stimulate DNA synthesis in human foreskin fibroblasts. Recombinant human bFGF 1-146 and [Ala69,Ser87]bFGF, an analogue where two of the four cysteines had been replaced by alanine and serine, were equipotent to standard bovine basic fibroblast growth factor. Substitution of aspartic acid-19 by arginine in the first heparin-binding domain yielded a molecule that stimulated a higher total mitogenic response in fibroblasts as compared to bFGF. In addition, replacement of either arginine-107 in the second domain or glutamine-123 in the third domain with glutamic acid resulted in compounds that were 2 and 4 times more potent than bFGF. In contrast, substitution of arginine-107 with isoleucine reduced the activity of the molecule by 100-fold. Combination of domain substitutions to generate the [Glu107,123]bFGF and [Arg19,Lys123,126]bFGF mutants did not show any additivity of the mutations on biological activity. Alterations in the biological activity of the analogues was dependent on both the site of and the type of modification. Increased positive charge in the first domain and increased negative charge in the second and third domains enhanced biological potency. The altered activities of the derivatives appear to be due in part to changes in the affinity of the analogues for heparin. We conclude that changes in all three of the putative heparin-binding domains result in altered mitogenic activity and heparin interaction of basic fibroblast growth factor.

Cyanophora paradoxa is a flagellated protozoan which possesses unusual, chloroplast-like organelles referred to as cyanelles. The psbE and psbF genes, which encode the two apoprotein subunits of cytochrome b-559, have been cloned from the cyanelle genome of C. paradoxa. The complete nucleotide sequences of these genes and their flanking sequences were determined by the chain-termination, dideoxy method. The psbE gene is composed of 75 codons and predicts a polypeptide of 8462 Da that is seven to nine residues smaller than most other psbE gene products. The psbF gene consists of 43 codons and predicts a polypeptide of 4761 Da. Two open reading frames, whose sequences are highly conserved among cyanobacteria and numerous higher plants, were located in the nucleotide sequence downstream from the psbF gene. The first open reading frame, denoted psbI, is composed of 39 codons, while the second open reading frame, denoted psbJ, is composed of 41 codons. The predicted amino acid sequences of the psbI and psbJ gene products predict proteins of 5473 and 3973 Da respectively. These proteins are probably integral membrane proteins anchored in the membrane by a single, transmembrane alpha helix. The psbEFIJ genes are probably co-transcribed and constitute an operon as found for other organisms. Each of the four genes is preceded by a polypurine sequence which resembles the consensus ribsosome binding sequences for Escherichia coli.

No abstract