| Phycobiliprotein are important light-harvesting pigment proteins in Cyanophyceae, Rhodophyceae, Cryptophyceae and a little Pyrrophyceae. Phycobiliprotein are classified according to their composition and spectrum properties: phycocyanin (610-640nm), phycoerythrin(500-570nm) and allophycocyanin (650-671nm). These proteins are not only used as industrial materials and food additives, but also found that they can cure some special diseases and so they are deserved attention and widely used in food industry, chemical industry, medical diagnose and clinical medicine, etc. In addition, phycobiliprotein as florescence probe are also deserved more and more attention because of their strong florescence, high quantum yields and easy combination with isotope and antibody.Phycobiliprotein are mainly prepared from Spirulina by isolation and purification. But the culture of the Spirulina is difficult and the large-scale culture of the Spirulina in outdoor condition lacks comprehensive research, so the above method has disadvantages: complex process, low yields and heavy material waste. It is essential to produce these proteins by genetic engineering.This dissertation includes the following sections:(1) Cloning and sequencing of the phycocyanin gene from Spirulina maxima and its evolutionary analysis. The results indicated that the length of the phycocyanin gene obtained is 1119bp and the homology with that of the phycocyanin gene from Spirulinaplatensis is 99% at the nucleotide acid level, β-subunit is upstream from a-subunit, with 111bp segment separating them. The α- and β-subunit gene-coding regions contain 489 and 519 bp, respectively. The a-subunit contains 162 amino acid residues, with methionine and serine at the NH2- and carboxyl-terminal amino acids, respectively. The β-subunit has 172 amino acid residues, with methionine and serine at the NH2- and carboxyl-terminal amino acids, respectively. The codons show asymmetries. In an examination for possible ribosome binding sites, the sequenceg-a-g-a was found 8-11 bp upstream from the β-subunit gene. This sequence is similar, although not identical, to the consensus prokaryotic ribosome binding sequence g-g-a-g. There are three chromophore-binding sites within the phycocyanin subunits, α-amino acid 84 and p-amino acids 82 and 153. The site at cysteine-153 is near the COOH terminus. It has been proposed that an insertion event, whereby a short stretch of 10 or 12 amino acids was introduced into the ancestral phycocyanin P-subunit gene product near its COOH terminus, could have generated this second chromophore-binding site. According to the model of Troxler et al, this second site in the P-subunit is due to the insertion of 12 amino acids at positions 146-157. The phycocyanin contains many hydrophobic amino acid residues and they play an important role in the aggregation of the phycocyanin. This shows that the phycocyanin may be produced by the replication of a hydrophobic protein. The homologies between a- and P- subunit of the phycocyanin, between α-subunit of the phycocyanin and a-subunit of the allophycocyanin, between P-subunit of the phycocyanin and P-subunit of the allophycocyanin, and between oc- and p- subunit of the allophycocyanin were also compared at the amino acid level. The homologies among them are arranged as follows: Hp_p> Hp.a> Ha.p> Ha-a. The differences in amino acid sequences between p hycocyanin and a llophycocyanin show that these proteins function differently in the transfer of the phycobilisome energy and the aggregation of these proteins, but the homology among these proteins also shows that the energy transfer function among phycobilisome must be similar. It is suggested that the genes for the allophycocyanin subunits arose first, and then gave rise to the genes for the phycocyanin subunits. The homology is higher between t he p hycocyanin β-subunit and the allophycocyanin than that between a- and p-subunit of the phycocyanin and that between a- and p-subunit of the allophycocyanin, which indicated that the gene r...
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