HETEROCYST - Cyanobacteria - page 232

exhibit considerable degree of similarity in the sequence of amino acids, three-dimensional structure
and other enzymatic properties with nitrogenases from Azotobacter vinelandii or Klebsiella pneumoniae
(Wolk et al ., 1994). There are two other proteins synthesized during nif gene expression in two
different cyanobacteria. The fi rst one is in Nostoc commune that synthesizes a hemoprotein, encoded
by glbN gene, located in between nifU and nifH , during prolonged incubation under microaerophilic
diazotrophic conditions (Potts et al ., 1992). The second one is in case of Anabaena sp. strain PCC 7120,
GroEL, a multifunctional chaperonin, is synthesized in vegetative cells and heterocysts during nif
gene expression (Jäger and Bergman, 1990) that probably helps in proper folding of nitrogenase
(Govezensky et al ., 1991).
The reductive nature of heterocysts has been highlighted by their ability to reduce 2,3,5-triphenyl
tetrazolium chloride (TTC; 0.01% w/v) and the red formazan crystals can be readily seen in the
heterocysts of A . variabilis , C . licheniforme , Nostoc sp. Calothrix sp., and Scytonema sp. under the light
microscope (Drawert and Metzner, 1956; Tischer, 1957). Their reductive nature and nitrogenase
activity in heterocysts has further been confi rmed (Talpasayi and Kale, 1967; Fay et al ., 1968; Stewart
et al ., 1969). A gradient in the formation of formazan in heterocysts and the intervening vegetative
cells does not result after treatment with TTC due to competition for electrons between TTC and O 2 .
On the other hand, treatment with 3,3',-(3,3'-dimethoxy-4,4'-biphenylene)-bis-(2-p-nitrophenyl)-5-
phenyl-2H tetrazolium chloride (NBT; 0.01% w/v) resulted in a gradient of the blue colour formation
highest (reducing power) in heterocysts and proheterocysts (of A . cylindrica , A . inaequalis , A . variabilis
and Anabaenopsis circularis ) followed by vegetative cells. Nitrogen fi xation by the heterocysts has
been found to be adversely affected by the reduction of TTC or NBT as well as the detachment
of heterocysts with the adjacent vegetative cells (Fay and Kulasooriya, 1972). It means that the
attachment of heterocysts with the adjacent vegetative cells is important for nitrogen fi xation to
proceed. In this connection, the results of Weare and Benemann (1973) are important as breakage of
fi laments by blending at the junctions of heterocysts with vegetative cells caused suffi cient decrease
in the capacity of nitrogen fi xation by A . cylindrica . Direct demonstration that heterocysts fi x nitrogen
came from studies on the incorporation of 15 N (Ohmori and Hattori, 1971) and 13 N (Wolk et al .,
1974) by the intact fi laments of A . cylindrica . Inhibition of 13 N fi xation into heterocysts was caused
by carbon monoxide, hydrogen gas or prior growth of the cyanobacterium in media enriched with
combined nitrogen. The principal products of 13 N-fi xation in a number of cyanobacteria examined ( A .
cylindrica , A . variabilis ATCC 29413, C . licheniforme ATCC 29412, Gloeothece sp. 6909, ATCC 27512 and
Plectonema boryanum UTCC 594) have been found to be [ 13 N]-NH 4 and [ 13 N]-glutamine (Thomas et
al ., 1977; Meeks et al. , 1978). A decrease in the label of glutamine and a corresponding increase in the
label in glutamate suggested the mediation of GS-GOGAT pathway for ammonia assimilation after
nitrogen fi xation or exogenous ammonia assimilation (Meeks et al ., 1978; Haselkorn, 1978). The above
observations establish clearly the role of heterocysts in nitrogen fi xation under aerobic conditions
but under specialized microaerophilic or anaerobic conditions the induction of nitrogenase in the
vegetative cells of A . cylindrica has also been demonstrated (Smith and Evans, 1970, 1971; Haystead
et al ., 1970; Van Gorkom and Donze, 1971). The ability of isolated heterocysts to perform nitrogen
fi xation has largely been shown by acetylene reduction assay (Stewart et al ., 1968).
Isolated heterocysts of A . cylindrica Lemm or A . variabilis ATCC 29413 reduced acetylene very
much like the intact fi laments in light and did not require any additional co-factors (Wolk and
Wojciuch, 1971a,b; Peterson and Burris, 1976; Jensen et al ., 1986). The requirement of light for
nitrogen fi xation by the isolated heterocysts of A . cylindrica and its inhibition by O 2 concentrations
greater than atmospheric levels and by dark exposure all go in favour of the heterocyst being the
site for nitrogen fi xation (Weare and Banemann, 1973). There are evidences for the synthesis of