Localization of glutamine synthetase in thin sections of nitrogen-fixing Anabaena cylindrica was performed using immuno-gold/transmission electronmicroscopy. The enzyme was present in all of the three cell types possible; vegetative cells, heterocysts and akinetes. The specific gold label was always more pronounced in heterocysts compared with vegetative cells, and showed a uniform distribution in all three types. No specific label was associated with subcellular inclusions such as carboxysomes, cyanophycin granules and polyphosphate granules. When anti-glutamine synthetase antiserum was omitted, no label was observed.

1989. Glycolate tnetabolism in cyanobacteria. 1. Glycolate excretion and phosphoglycolate phosphatase activity. -Physiol. Plant. 75: 137-143.A comparative analysis of glycolate excretion in 11 cyanobacteria showed that 8 strains, although grown and assayed in air, excreted glycolate. The largest quantities were excreted by the filamentous strains Plectonema boryanum 73110 and Anabaena cylindrica (Lemm). The carbon lost by excretion was at most 9% of the net fixed carbon in air for heterocystous cyanobacteria but increased (up to 60%) in some strains under a high pO, (0.03 kPa CO, in pure O-,). ^. cylindrica excreted glycolate at a maximum level of 2 and 10 \imo\ (mg chl a)"' h"' in air and at high pOj, respectively. The excretion continued for several hours. Increases in light intensity and pO2 and a shift in pH from 7 to 9 increased the amount of glycolate excreted. A. cylindrica also showed the most Oi-sensitive fixation of COi. In vitro activity of phosphoglycolate phosphatase (EC 3.1.3.18) was found in all strains tested, with the highest activities noted for Gloeobacter violaceus 7.82 and Gloeothece 6909 and for young cultures oi A, cylindrica. The lowest activities were found in Anabaena 7120 and Anacystis nidulans 625, strains excreting no or only minor quantities of glycolate.

Aminooxyacetate and aminoacetonitrile cause increased excretion of glycolate by the cyanobacterium Anabaena cylindrica. Both compounds also reduce NH4-N release induced by methionine sulfoximine in nonnitrogen-fixing cultures. Changes in amino acid pool sizes together with changes in activities of some enzymes related to glycolate metabolism show that glyoxylate to glycine conversion and glycine to serine conversion are inhibited by aminooxyacetate and aminoacetonitrile, respectively. The results also verify that photorespiratory glycolate metabolism via amination of glyoxylate is operative in A. cylindrica.We have recently reported on the stimulation of glycolate excretion by INH2 in cultures of the cyanobacterium Anabaena cylindrica (4). INH inhibits the conversion of glycine to serine in the glycolate pathway of higher plants and eukaryotic algae (27). Evidence has also been presented for the photorespiratory production of ammonia by A. cylindrica (3). As with some green algae (13,23) Medium and Cultivation. Cells were grown in pure culture in either NH4Cl-supplemented (N+) or N-free (N-) BG 11 medium (26) and sparged continuously with sterile air. Cultivation and cell harvesting from N+ and N-media were carried out as detailed earlier (1, 3, 4). The N+-grown cultures were used in experiments on the release of ammonium and amino acid pool sizes, while N-grown cells were used in the remainder of the experiments. Solutions ofall inhibitors were adjusted to pH 7.5 before addition to the medium.Glycolate Excretion. The procedure given earlier (4) was used and excreted glycolate assayed according to Calkins (6). The cultures were grown on air before use and then sparged with 100% 02 from the beginning of the excretion experiments. A photon fluence rate of 200 Mumol. m2 . s' at the surface of the vessels was used during excretion assays, provided by white fluorescent tubes.Release of NH4-N. MSX (0.2 mM)-induced release of NH4-N to the medium was determined as detailed earlier (3), using the phenol hypochlorite method of Chaney and Marbach (7). Cultures were pretreated only with AOA, AAN, and INH as they all, as such, interfere with the colorimetric assay by producing a blue color similar to that of indophenol blue. Cells were centrifuged after exposure to the inhibitors, carefully washed, and resuspended in fresh N-free medium containing 0.2 mM MSX (time zero). The release of NH4-N to the medium was then determined at the time intervals given in Tables III and IV

1989. Glycolate metabolism in cyanobacteria. 11. Evidence for a mediated transport of glycolate in Anabaena 7120. -Physiol. Plant. 75;[144][145][146][147][148][149][150] The mechanism for uptake of glycolate in the cyanobacterium Anabaena 7120 and its capacity to metabolize glycolate were examined. The uptake of ['^C]-glycolate in light, at pH 7, consisted of^ an initial rapid phase (<60 s) and a second slower phase. The latter obviously represents metabolism as the glycolate dehydrogenase inhibitor 2-pyridylhydroxymethanesulfonic acid (HPMS) did not affect the initial uptake phase while the second phase was strongly reduced. The sulfhydryl reagent N-ethylmaleimide (NEM) inhibited uptake of glycolate and the uptake was reduced by lactate, glycerate and glyoxylate. Treatment with triphenylmethylphosphonium (TPMP^), a lipophilic cation collapsing A\p only slightly reduced the uptake of glycolate. At pH 7.0, the F,,F,-ATPase inhibitor N,N'-dieyclohexylcarbodiimide (DCCD) and the uncoupler carbonylcyanide m-chlorophenylhydrazone (CCCP) abolished the uptake. Inhibition of photophosphorylation by dark-treatment and presence of 3-(3',4'-dichlorophenyl)-l,l-dimethylurea (DCMU) also reduced the uptake. Decreasing the pH in the range of 10 to 5.5 increased the uptake. In contrast to the situation at pH 7, CCCP did not affect the initial glycolate uptake at pH 5.5. We conclude that the uptake of glycolate is a carrier-mediated process which, at pH 7, is dependent on a H^-ATPase to create the ApH across the membranes needed for uptake, while at pH 5.5 the uptake of glycolate is not ATP-dependent. The capacity to metabolize glycolate was at least 50 (xmol (mg chl a)"' h"' in young cultures. In older cultures the rate was nearly 50% lower.