Isolation and overexpression in Escherichia coli of the flavodoxin gene from Anabaena PCC 7119

Abstract

The gene coding for flavodoxin from Anabaena PCC 7119 was cloned by using the polymerase chain reaction (PCR). The gene is transcribed into a 1250-base transcript. The expression of the flavodoxin gene was analysed and found to be regulated at the transcriptional level by the availability of iron. The PCR-amplified gene was cloned into the expression vector pTrc 99b and expressed in Escherichia coli. High concentrations of flavodoxin were found (20% of total protein). The recombinant protein was purified from the cytosolic fraction of the cells and it exhibited properties identical with those of the wild-type Anabaena flavodoxin.

A laser flash absorption spectroscopy study of Anabaena sp. PCC 7119 flavodoxin photoreduction by photosystem I particles from spinach

“…Since the gene of Anabaena sp. PCC 7119 Fld has been cloned and over-expressed in E. coil [23], and work on the preparation of Fld mutants is in progress, using the methods described here it will be possible to determine whether or not the same region of the Fld molecule interacts with both PSI and FNR. Furthermore, a comparative study of PSI reduction of Fd and Fld mutants will help to elucidate the factors which control these electron transfer processes.…”

Interaction between Photosystem I and Flavodoxin from the Cyanobacterium Synechococcus sp. PCC 7002 as Revealed by Chemical Cross‐Linking

“…PCC 7002, essentially as described previously (Golbeck et al, 1988). Recombinant flavodoxin of the same organism was expressed in Escherichia coli strain BL21 harboring isiB (Leonhardt and Straus, 1992) in the expression vector pSE280 (Brosius, 1989) and purified esseiitially as dcscribed (Fillat et al, 1991). Ferredoxin from Synechococccis sp.…”

Conformational stability of apoflavodoxin

“…I) as a model protein for stability and folding studies. This protein appears well suited for the purpose: its crystal structure is known (Rao et al, 1992, for the holo form; Genzor et al, 1996, for the apo form), its gene has been cloned (Fillat et al, 1991), and the protein can be expressed in Escherichia coli with good yields. Flavodoxin contains four tryptophane residues, one trans proline, and no disulphide bridges.…”