Immunoaffinity Purification and Characterization of Cyclodextrin Glycosyltransferase from Bacillus circulans A11

ABSTRACT The purified IgG fraction of anti-CGTase was coupled to CNBr-activated Sepharose 4B and used as immunoaffinity gel to purify CGTase from Bacillus circulans A11. The enzyme was successfully purified to approximately 155 folds with a 45% yield and specific activity of 3302 units/mg protein. It was a single polypeptide of 72 KDa. The amino acid composition of this CGTase was found to contain high amounts of Asx, Glx, Gly, Ala, and Thr and low amounts of His, Met, Cys, and Trp. N-Terminal sequence was A P D T S V S N K Q N F S T D V I Y Q I. Chemical modification and substrate protection studies indicate the presence of Trp, His, Tyr, and Asx/Glx residues at the active site of CGTase, while Cys, Lys, and Ser were proved to have no influence on CGTase catalysis. From HPLC analysis of products of enzymatic reaction, this CGTase produced mainly β-CD. The ratio of α : β : γ -CD was 1 : 4.1 : 1.1. When analyzed by non-denaturing PAGE, the enzyme demonstrates the isoform pattern. Four isoforms with the same molecular mass but different in quantity and pI values were observed. All isoforms demonstrate amylolytic and cyclizing activities of CGTase

Genomic Structure of an Economically Important Cyanobacterium, Arthrospira (Spirulina) platensis NIES-39

A filamentous non-N2-fixing cyanobacterium, Arthrospira (Spirulina) platensis, is an important organism for industrial applications and as a food supply. Almost the complete genome of A. platensis NIES-39 was determined in this study. The genome structure of A. platensis is estimated to be a single, circular chromosome of 6.8 Mb, based on optical mapping. Annotation of this 6.7 Mb sequence yielded 6630 protein-coding genes as well as two sets of rRNA genes and 40 tRNA genes. Of the protein-coding genes, 78% are similar to those of other organisms; the remaining 22% are currently unknown. A total 612 kb of the genome comprise group II introns, insertion sequences and some repetitive elements. Group I introns are located in a protein-coding region. Abundant restriction-modification systems were determined. Unique features in the gene composition were noted, particularly in a large number of genes for adenylate cyclase and haemolysin-like Ca2+-binding proteins and in chemotaxis proteins. Filament-specific genes were highlighted by comparative genomic analysis