HETEROCYST - Cyanobacteria - page 174

synthesis is mediated by a non-ribosomal synthetic process (Simon, 1973b) and the basic requirements
of the biosynthetic reaction have been studied in vitro that required the two constituent amino acids,
Mg-ATP, cyanophycin primer and the enzyme (Simon, 1976). Cultures of Aphanocapsa sp. 6308
(ATCC 27150) starved of light, CO 2 , sulphur and phosphorus showed enhanced content of CGP
when compared to control cultures. However, nitrogen limitation or lowering of growth temperature
caused a decrease in CGP content (Allen et al ., 1980). A number of studies have indicated that the
CGP in cyanobacterial cells is broken down during nitrogen deprivation and the synthesis of CGP
ensues once nitrogen is replenished in the medium (Allen and Hutchinson, 1980; Allen and Hawley,
1983; Mackerras et al ., 1990). In view of this, it is suggested that CGP acts as a transient reservoir of
nitrogenous material (Shively, 1974; Carr, 1988; Mackerras et al ., 1990; Li et al ., 2001). The synthesis
of CGP depended on the degradation of cellular proteins as well as newly assimilated nitrogen in
A . cylindrica (Simon, 1973b) and Aphanocapsa sp. 6308 (ATCC 27150) (Allen and Hawley, 1983). This
has been confi rmed in Synechocystis sp. strain PCC 6803 by the use of N 15 labelling and H 1 NMR
spectroscopy (Allen et al ., 2005) and proton nuclear magnetic resonance spectroscopy (Kolodny et
al ., 2006). The synthesis of CGP took place by the addition of monomers to the pre-existing polymer
(Allen et al ., 2005). The non-template-dependent polymerization of CGP takes place in three steps
of chain initiation, elongation and termination mediated by cyanophycin synthetase (CphA).
The enzyme fom A . variabilis ATCC 29413 (Ziegler et al ., 1998), Synechocystis sp. strain PCC 6803
(Aboulmagd et al ., 2000, 2001) and the thermophilic Synechococcus sp. strain MA19 (Hai et al ., 1999,
2002) has been purifi ed. The specifi c activity of the enzyme from A . variabilis ATCC 29413 is 2 to 5
fold higher than that of the enzyme from Synechocystis sp. strain PCC 6803 and the cphA gene from
A . variabilis ATCC 29413 has been cloned and expressed in E . coli . CphA from A . variabilis ATCC
29413, with a molecular weight of 100 kD, appeared as a single band in sodium dodecyl sulphate/
polyacrylamide gel electrophoresis. Since the native enzyme has a molecular weight of 230 kDa,
determined as per size-exclusion chromatography, it is suggested that the native enzyme exists as
a dimer with two ATP-binding sites (Zeigler et al ., 1998). The cphA gene from Anabaena sp. strain
PCC 7120 has been expressed in E . coli and the recombinant protein exhibited highest specifi c CphA
activity by synthesizing CGP expressed as 6.7 nmol arginine min -1 mg protein -1 (Voss et al ., 2004).
Gupta and Carr (1981b) have found that the enzymes involved in the synthesis ( cphA ) and
breakdown (cyanphycinase, cphB ) of CGP are more active in heterocysts than vegetative cells. The
accumulation of higher levels of CGP in the heterocysts signifi es that it may serve as a reserve
material for biosynthetic reactions (Carr, 1988). The expression of nifH (nitrogenase protein) at the
late stage of heterocyst differentiation in Anabaena sp. strain PCC 7120 is followed by the synthesis
of CGP and its accumulation at the neck joining the heterocyst with the vegetative cells fi xes the
formation of polar nodule (Simon, 1987; Sherman et al ., 2000). During nitrogen defi ciency, there is
rapid mobilization of CGP in A . cylindrica and Synechocystis sp. strain 6803 (Mackerras et al ., 1990).
The diazotrophic unicellular Cyanothece sp. ATCC 51142 showed accumulation of CGP after a peak in
nitrogenase synthesis and the knock-out mutants (of cphA and cphB ) of this organism had to depend
on phycobilisome degradation in the absence of CGP synthesis (Li et al ., 2001). The growth properties
of a mutant of A . variabilis ATCC 29413, isolated after insertional mutagenesis of cphA gene, revealed
the non-essential nature of CGP metabolism for aerobic nitrogen fi xation. The mutant in the absence
of CGP synthesis and the polar nodule in the heterocysts exhibited a similar growth behaviour as the
wild-type irrespective of the nitrogen source and fi xed nitrogen under aerobic conditions (Ziegler et
al ., 2001). Two gene clusters, cph1 and cph2 governing the synthesis and breakdown of CGP in the
genome of Anabaena sp. strain PCC 7120 have been identifi ed. The expression of cph1 ( cphA1 and
cphB1 ) and cph2 ( cphA2 and cphB2 ) gene clusters although was noted in ammonium, nitrate and