Proc. Nati. Acad. Scd. USA Vol. 76, No. 1, pp. *265-267p January 1979
Biochemistry
Evidence for tetradecanal as the natural aldehyde in bacterial bioluminescence (fatty acids/luciferase/bacterial luminescence)
S. ULITZUR* AND J. W. HASTINGSt *Department of Food Engineering and Biotechnology, Technion-Israel Institute for Technology, Haifa, Israel; and tThe Biological Laboratories, Harvard
University, Cambridge, Massachusetts 02138
Communicated by W. D. McElroy, November 6,1978
ABSTRACT Dim aldehyde mutants of the luminous bacterium Beneckea harveyi emit light with exogenously added long-chain aliphatic aldehyde. In one class of these mutants, luminescence is also stimulated by myristic (tetradecanoic) acid. In such mutants the amount of light obtained by the addition of a small (limiting) amount of either tetradecanal or myristic acid may be increased 6fold by cyanide and other agents that block respiration. This indicates that the fatty acid product of the luminescent reaction is recycled. The effect, like the stimulation by exogenous fatty acid, exhibits specificity for the 14-carbon compound, suggesting that tetradecanal is the natural aldehyde. In those aldehyde mutants that are not stimulated to emit light by fatty acids, and thus presumably lack the recycling system, the chain-length-specific stimulation by cyanide does not occur.
MATERIALS AND METHODS Mutants of B. harveyi 392 (9) were obtained after mutagenesis with nitrosoguanidine, as described (6). The mutants used were (i) M17, a dim mutant that emits light when exposed to either exogenous aldehyde or myristic acid (6, 8); (ii) M42, a dim mutant isolated in this study that emits light only with added aldehyde; and (iii) TSAS-F1, a temperature-conditional mutant (normal luminescence at 250C) that is dim at 360C but emits light only with added aldehyde (6, 8). Cells were grown at the temperature specified, with shaking, in a complex medium consisting of artificial sea water and 20 mM 4-morpholinepropanesulfonic acid (Mops, Sigma) buffer (pH 7.3) to which 0.5% peptone (Difco), 0.3% yeast extract (Difco), and 0.3% glycerol were added. Artificial sea water contains (g/liter): NaCl, 17.55; KCl, 0.75; MgSO4-H20, 12.3; CaCl2-2H20, 1.45; and K2HPO4-3H20, 0.075. Cell density was determined in a Klett-Summerson photometer (filter number 66); 100 Klett units was equal to about 5 X 108 cells ml-'. Bioluminescence was measured in a photomultiplier photometer (10) and expressed in quanta sec-1 (11). Fatty acids, aldehydes, and 2heptyl-4-hydroxyquinoline-N-oxide (HQNO) (all >99% pure; Sigma) were dissolved in absolute ethanol. Sodium azide (NaN3) was dissolved in water and 0.1 M KCN was prepared fresh each day in 20 mM Mops (pH 7.5). The oxygen uptake of a growing cell culture at pH 7.5, as well as washed cell suspensions at pH 6.5, as measured with an oxygen electrode (12), was inhibited by >99% by 1 mM cyanide, 5 mM NaN3, or 10,gM HQNO.
The involvement of aldehyde in bacterial bioluminescence has been known for many years (1), but neither its metabolism nor the chemical identity of the cellular aldehyde has been elucidated (2, 3). The in vitro reaction involves the mixed function oxidation of FMNH2 and a long-chain aliphatic aldehyde to give the corresponding acid and light emission. Pure Beneckea harveyi luciferase is not highly specific with regard to aldehyde chain length (8-14 carbons), even though. some distinctions can be made, especially with luciferases from different bacterial species (3, 4). The involvement of aldehyde is also known in vivo from dim aldehyde mutants, which emit light at near-normal levels upon exposure to long-chain aldehyde (5, 6). Here, also, the bioluminescence response occurs with aldehydes of different chain lengths (7, 8). However, neither the in vitro nor the in vivo studies have led to the identification of the "natural" aldehyde. Recently it was found that some (but not all) aldehyde mutants respond also to exogenous long-chain fatty acid (8). However, these mutants exhibit specificity for myristic (tetradecanoic) acid, suggesting that this chain length may be the natural aldehyde. Since fatty acid is the postulated product in the luciferase reaction, recycling of the product with high specificity for added tetradecanal would be expected in such mutants. This paper shows that this occurs and, further, that, under conditions where respiration is blocked and highly reducing intracellular conditions are expected, the quantum yields of both tetradecanal and myristic acid are increased as much as 60-fold.
RESULTS AND DISCUSSION As shown in Table 1 the in vivo responses of M17 cells to aldehydes of different even-numbered (the naturally occurring) chain length, ranging from 8 to 16 carbons, are similar within a factor of about 4. Responses within this range were also demonstrated in vitro for purified B. harveyi luciferase (4). We considered, therefore, that the response in vivo might be interpreted simply as a reflection of the activity of cellular luciferase with the different aldehydes. However, if recycling of a certain aldehyde occurs, then an increase in photon yield from such an aldehyde should occur under conditions more favorable for recycling. Cyanide, like other respiratory inhibitors and low oxygen, may be expected to result in an increase in the steady-state intracellular level of reduced coenzymes, such as NADH and FMNH2, which might
The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.
Abbreviations: Mops, 4-morpholinepropanesulfonic acid; HQNO, 2-heptyl-4-hydroxyquinoline-N-oxide. 265
Biochemistry: Ulitzur and Hastings
266
Chain
length
-CN-
8 10 12 14 16 None
8.0 20.0 5.0 9.0 8.0 0.1
Proc. Nati. Acad. sci. USA 76 (1979)
Table 1. Effect of cyanide on photon yields in aldehyde mutants Aldehyde added M17 M42 TSAS-F1 +CN-CN+CN-CN+CN13.0 45.0 20.0 600.0 17.0 0.5
8.0 30.0 4.0 2.0 2.0 0.04
15.0 60.0 6.0 4.0 3.0 0.1
9.0
40.0
0.2
0.5
0.02
0.06
Acid added M17 -CN+CN1.0 1.3 1.1 1.0 4.0 11.0 150.0 2400.0 1.4 6.0 0.1 0.5
B. harveyi aldehyde mutants M17 and M42 were grown at 300C to a density of about 1 X 109 cells ml-', harvested by centrifugation, and resuspended in buffered artificial sea water at pH 6.5 at a final cell density of 2 X 108 cells ml-'. For mutant TSAS-F1, the procedures were similar except the cells were grown and measured without harvesting (i.e., directly in the growth medium) at 360C. Only decanal and tetradecanal were tested with this mutant. One nanomole (6 X 1014 molecules) of each of the acids or the aldehydes was added and the resulting luminescence at 250C was recorded and integrated with and without added cyanide (1 mM). Light emission is expressed in photons; the values given should be multiplied by 1011. The bottom line (None) shows photon yields with and without added cyanide, but with no aldehyde or acid added. These values, though always low, are sometimes compromised by the presence of contaminating levels of aldehyde or fatty acids in the vials.
thereby result in an increase in the luminescence if FMNH2 is limiting in vivo. However, given the appropriate enzymatic pathways, the presence of cyanide could also result in an increase in the rate of reduction of the long-chain fatty acid to aldehyde and thus enhance turnover or recycling of the longchain compound. When cyanide (1 mM) was added, an increase (2-4 times) in the total light emitted by M17 cells occurred with most of the aldehydes (Table 1). This in itself might simply reflect an effect of cyanide on the level of reduced FMN. With tetradecanal, however, the increase was far more (60 times), corresponding to a yield of about 0.1 photon per molecule added. Since the bioluminescent quantum yield in the in vitro reaction is estimated to be about 0.1 (3), this result is compatible with the possibility that all the added aldehyde molecules are used. Because some of the added aldehyde may be expected to be lost via autooxidation and cytoplasmic reactions with amino and sulfhydryl groups of proteins (13), this yield may be con4
sidered to be a minimum. It may thus be inferred that some turnover (recycling) of the added aldehyde is occurring, even if the quantum yield in vivo were somewhat higher than 0.1.
Another class of aldehyde mutants responds to aldehydes but not to any of the fatty acids. These mutants, which are deduced to be defective in the step that converts myristic acid to tetradecanal, should thus also be unable to use fatty acid formed as a product of bioluminescence and, therefore, would not exhibit turnover of added aldehyde. Results with two such mutants, M42 and TSAS-F1, demonstrate that this is so (Table 1). The effect of cyanide upon the photon yield of both mutants with added aldehyde again shows an increase with cyanide that is relatively small but about the same with aldehydes of different chain lengths. This result is similarly attributed to the higher level of FMNH2 in the cyanide-treated cells and thus shows that cyanide does not act simply.by enhancing aldehyde utilization by luciferase.
-
0
3 2 C~~~~~~~~~~~~
o
2
0
4
6
10
C
600 800 1000 Time, min FIG. 1. B. harveyi M17 cells were grown at 3000 to a density of 1 X 109 cells ml-', washed once by centrifugation, and suspended in artificial sea water/Mops, pH 6.5 at a density of 5 X 107 cells ml-'. One milliliter of culture was placed in a scintillation vial and 25 pmol (1.5 X 1013 molecules) of myristic acid was added without (Inset) or in the presence of 2.5 mM KCN at 220C. The light was recorded and integrated until the reaction was essentially complete. The background luminescence was determined and subtracted. Luminescence is in light units; 1 unit = 108 quanta-sec'1. 0
200
400
0
25
50
75
Time, min FIG. 2. M17 cells were grown, washed, and resuspended as described for Fig. 1. To each aliquot, 50 pmol of myristic acid was added. The luminescence was then recorded with no further additions (-), with added 1 mM KCN (o), 10,uM HQNO (A), or 5 mM sodium azide (o), or in equilibrium with an atmosphere of 0.2% 02 in N2 (o). Also shown is the effect the secondary addition of 1 mM cyanide (X) and 10 MM HQNO (A) during the luminescence decay. Luminescence is in light units; one unit = 1.7 X 108 quanta .isec-1.
Proc. Natl. Acad. Sci. USA 76 (1979)
Biochemistry: Ulitzur and Hastings The stimulatory effect of cyanide is not confined to the 14-carbon aldehyde. With the M17 mutant there is a large increase in the quantum yield of myristic acid in the presence of cyanide (Fig. 1), much more than with acids of other chain lengths (Table 1). The absolute photon yield with added tetradecanoic acid was always higher than with tetradecanal; quantum yields between 0.5 and 1 were commonly obtained with the acid when respiration was blocked. The reasons for this better yield may include solubility factors and the occurrence of so many competing side reactions for the aldehyde (13). With both acid and aldehyde (14 carbons) a similar stimulatory effect upon photon yields with mutant M17 occurs with other inhibitors of electron transport (HQNO and sodium azide) as well as under low (t0.2%) 02 (Fig. 2). These results indicate that the natural endogenous aldehyde in bacterial bioluminescence is the 14-carbon, saturated, aliphatic compound, and that there are cellular enzymes able to recycle its oxidation product (but not other chain lengths) by reduction. Although the results presented are for B. harveyi, similar observations in vivo indicate that the conclusions are also applicable for Photobacterium fischeri (8) and P. leiognathi. The small quantities of aldehyde that could be isolated from two species of luminous bacteria were mostly of chain lengths 12, 14, and 16 carbons, in relative abundance of 5, 63, and 30%, respectively, in one species and 36, 32, and 20% in the other (14). It is also interesting that with luciferase isolated from all three Photobacterium species the rate of the reaction is much greater with tetradecanal than with aldehydes of other
267
chain lengths, especially the naturally occurring even-numbered ones (ref. 15; unpublished data). 1. Strehler, B. L. & Cormier, J. J. (1953) Arch. Biochem. Biophys.
47,916-3.
2. Hastings, J. W. (1978) Photochem. Photobiol. 27, 397-404. 3. Hastings, J. W. & Nealson, K. H. (1977) Annu. Rev. Microbiol.
31,549-595. 4. Hastings, J. W., Weber, K., Friedland, J., Eberhard, A., Mitchell, F. W. & Gunsalus, A. (1969) Biochemistry 8, 4681-4689. 5. Rogers, P. & McElroy, W. D. (1955) Proc. Natl. Acad. Sci. USA 41, 67-70. 6. Cline, T. W. & Hastings, J. W. (1971) Proc. Natl. Acad. Sci. USA 68, 500-504. 7. Rogers, P. & McElroy, W. D. (1958) Arch. Biochem. Biophys.
75,87-105.
8. Ulitzur, S. & Hastings, J. W. (1978) Proc. Natl. Acad. Sci. USA
75,266-269.
9. Reichelt, J. L. & Baumann, P. (1973) Arch. Mikrobiol. 94,
283-330.
10. Mitchell, G. & Hastings, J. W. (1971) Anal. Biochem. 39, 243250. 11. Hastings, J. W. & Weber, G. (1963) J. Opt. Soc. Am. 53, 1410-1415. 12. Ulitzur, S. & Hastings, J. W. (1978) J. Bacteriol. 133, 13071313. 13. Schauenstein, H., Esterbauer, H. & Zollner, H. (1977) Aldehydes in Biological Systems (Pion, London). 14. Shimomura, O., Johnson, F. H. & Morise, H. (1974) Proc. NatI. Acad. Sci. USA 71, 4666-4669. 15. Hastings, J. W., Spudich, J. A. & Malnic, G. (1963) J. Biol. Chem.
a238,3100-3105.
Biochemistry
Evidence for tetradecanal as the natural aldehyde in bacterial bioluminescence (fatty acids/luciferase/bacterial luminescence)
S. ULITZUR* AND J. W. HASTINGSt *Department of Food Engineering and Biotechnology, Technion-Israel Institute for Technology, Haifa, Israel; and tThe Biological Laboratories, Harvard
University, Cambridge, Massachusetts 02138
Communicated by W. D. McElroy, November 6,1978
ABSTRACT Dim aldehyde mutants of the luminous bacterium Beneckea harveyi emit light with exogenously added long-chain aliphatic aldehyde. In one class of these mutants, luminescence is also stimulated by myristic (tetradecanoic) acid. In such mutants the amount of light obtained by the addition of a small (limiting) amount of either tetradecanal or myristic acid may be increased 6fold by cyanide and other agents that block respiration. This indicates that the fatty acid product of the luminescent reaction is recycled. The effect, like the stimulation by exogenous fatty acid, exhibits specificity for the 14-carbon compound, suggesting that tetradecanal is the natural aldehyde. In those aldehyde mutants that are not stimulated to emit light by fatty acids, and thus presumably lack the recycling system, the chain-length-specific stimulation by cyanide does not occur.
MATERIALS AND METHODS Mutants of B. harveyi 392 (9) were obtained after mutagenesis with nitrosoguanidine, as described (6). The mutants used were (i) M17, a dim mutant that emits light when exposed to either exogenous aldehyde or myristic acid (6, 8); (ii) M42, a dim mutant isolated in this study that emits light only with added aldehyde; and (iii) TSAS-F1, a temperature-conditional mutant (normal luminescence at 250C) that is dim at 360C but emits light only with added aldehyde (6, 8). Cells were grown at the temperature specified, with shaking, in a complex medium consisting of artificial sea water and 20 mM 4-morpholinepropanesulfonic acid (Mops, Sigma) buffer (pH 7.3) to which 0.5% peptone (Difco), 0.3% yeast extract (Difco), and 0.3% glycerol were added. Artificial sea water contains (g/liter): NaCl, 17.55; KCl, 0.75; MgSO4-H20, 12.3; CaCl2-2H20, 1.45; and K2HPO4-3H20, 0.075. Cell density was determined in a Klett-Summerson photometer (filter number 66); 100 Klett units was equal to about 5 X 108 cells ml-'. Bioluminescence was measured in a photomultiplier photometer (10) and expressed in quanta sec-1 (11). Fatty acids, aldehydes, and 2heptyl-4-hydroxyquinoline-N-oxide (HQNO) (all >99% pure; Sigma) were dissolved in absolute ethanol. Sodium azide (NaN3) was dissolved in water and 0.1 M KCN was prepared fresh each day in 20 mM Mops (pH 7.5). The oxygen uptake of a growing cell culture at pH 7.5, as well as washed cell suspensions at pH 6.5, as measured with an oxygen electrode (12), was inhibited by >99% by 1 mM cyanide, 5 mM NaN3, or 10,gM HQNO.
The involvement of aldehyde in bacterial bioluminescence has been known for many years (1), but neither its metabolism nor the chemical identity of the cellular aldehyde has been elucidated (2, 3). The in vitro reaction involves the mixed function oxidation of FMNH2 and a long-chain aliphatic aldehyde to give the corresponding acid and light emission. Pure Beneckea harveyi luciferase is not highly specific with regard to aldehyde chain length (8-14 carbons), even though. some distinctions can be made, especially with luciferases from different bacterial species (3, 4). The involvement of aldehyde is also known in vivo from dim aldehyde mutants, which emit light at near-normal levels upon exposure to long-chain aldehyde (5, 6). Here, also, the bioluminescence response occurs with aldehydes of different chain lengths (7, 8). However, neither the in vitro nor the in vivo studies have led to the identification of the "natural" aldehyde. Recently it was found that some (but not all) aldehyde mutants respond also to exogenous long-chain fatty acid (8). However, these mutants exhibit specificity for myristic (tetradecanoic) acid, suggesting that this chain length may be the natural aldehyde. Since fatty acid is the postulated product in the luciferase reaction, recycling of the product with high specificity for added tetradecanal would be expected in such mutants. This paper shows that this occurs and, further, that, under conditions where respiration is blocked and highly reducing intracellular conditions are expected, the quantum yields of both tetradecanal and myristic acid are increased as much as 60-fold.
RESULTS AND DISCUSSION As shown in Table 1 the in vivo responses of M17 cells to aldehydes of different even-numbered (the naturally occurring) chain length, ranging from 8 to 16 carbons, are similar within a factor of about 4. Responses within this range were also demonstrated in vitro for purified B. harveyi luciferase (4). We considered, therefore, that the response in vivo might be interpreted simply as a reflection of the activity of cellular luciferase with the different aldehydes. However, if recycling of a certain aldehyde occurs, then an increase in photon yield from such an aldehyde should occur under conditions more favorable for recycling. Cyanide, like other respiratory inhibitors and low oxygen, may be expected to result in an increase in the steady-state intracellular level of reduced coenzymes, such as NADH and FMNH2, which might
The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.
Abbreviations: Mops, 4-morpholinepropanesulfonic acid; HQNO, 2-heptyl-4-hydroxyquinoline-N-oxide. 265
Biochemistry: Ulitzur and Hastings
266
Chain
length
-CN-
8 10 12 14 16 None
8.0 20.0 5.0 9.0 8.0 0.1
Proc. Nati. Acad. sci. USA 76 (1979)
Table 1. Effect of cyanide on photon yields in aldehyde mutants Aldehyde added M17 M42 TSAS-F1 +CN-CN+CN-CN+CN13.0 45.0 20.0 600.0 17.0 0.5
8.0 30.0 4.0 2.0 2.0 0.04
15.0 60.0 6.0 4.0 3.0 0.1
9.0
40.0
0.2
0.5
0.02
0.06
Acid added M17 -CN+CN1.0 1.3 1.1 1.0 4.0 11.0 150.0 2400.0 1.4 6.0 0.1 0.5
B. harveyi aldehyde mutants M17 and M42 were grown at 300C to a density of about 1 X 109 cells ml-', harvested by centrifugation, and resuspended in buffered artificial sea water at pH 6.5 at a final cell density of 2 X 108 cells ml-'. For mutant TSAS-F1, the procedures were similar except the cells were grown and measured without harvesting (i.e., directly in the growth medium) at 360C. Only decanal and tetradecanal were tested with this mutant. One nanomole (6 X 1014 molecules) of each of the acids or the aldehydes was added and the resulting luminescence at 250C was recorded and integrated with and without added cyanide (1 mM). Light emission is expressed in photons; the values given should be multiplied by 1011. The bottom line (None) shows photon yields with and without added cyanide, but with no aldehyde or acid added. These values, though always low, are sometimes compromised by the presence of contaminating levels of aldehyde or fatty acids in the vials.
thereby result in an increase in the luminescence if FMNH2 is limiting in vivo. However, given the appropriate enzymatic pathways, the presence of cyanide could also result in an increase in the rate of reduction of the long-chain fatty acid to aldehyde and thus enhance turnover or recycling of the longchain compound. When cyanide (1 mM) was added, an increase (2-4 times) in the total light emitted by M17 cells occurred with most of the aldehydes (Table 1). This in itself might simply reflect an effect of cyanide on the level of reduced FMN. With tetradecanal, however, the increase was far more (60 times), corresponding to a yield of about 0.1 photon per molecule added. Since the bioluminescent quantum yield in the in vitro reaction is estimated to be about 0.1 (3), this result is compatible with the possibility that all the added aldehyde molecules are used. Because some of the added aldehyde may be expected to be lost via autooxidation and cytoplasmic reactions with amino and sulfhydryl groups of proteins (13), this yield may be con4
sidered to be a minimum. It may thus be inferred that some turnover (recycling) of the added aldehyde is occurring, even if the quantum yield in vivo were somewhat higher than 0.1.
Another class of aldehyde mutants responds to aldehydes but not to any of the fatty acids. These mutants, which are deduced to be defective in the step that converts myristic acid to tetradecanal, should thus also be unable to use fatty acid formed as a product of bioluminescence and, therefore, would not exhibit turnover of added aldehyde. Results with two such mutants, M42 and TSAS-F1, demonstrate that this is so (Table 1). The effect of cyanide upon the photon yield of both mutants with added aldehyde again shows an increase with cyanide that is relatively small but about the same with aldehydes of different chain lengths. This result is similarly attributed to the higher level of FMNH2 in the cyanide-treated cells and thus shows that cyanide does not act simply.by enhancing aldehyde utilization by luciferase.
-
0
3 2 C~~~~~~~~~~~~
o
2
0
4
6
10
C
600 800 1000 Time, min FIG. 1. B. harveyi M17 cells were grown at 3000 to a density of 1 X 109 cells ml-', washed once by centrifugation, and suspended in artificial sea water/Mops, pH 6.5 at a density of 5 X 107 cells ml-'. One milliliter of culture was placed in a scintillation vial and 25 pmol (1.5 X 1013 molecules) of myristic acid was added without (Inset) or in the presence of 2.5 mM KCN at 220C. The light was recorded and integrated until the reaction was essentially complete. The background luminescence was determined and subtracted. Luminescence is in light units; 1 unit = 108 quanta-sec'1. 0
200
400
0
25
50
75
Time, min FIG. 2. M17 cells were grown, washed, and resuspended as described for Fig. 1. To each aliquot, 50 pmol of myristic acid was added. The luminescence was then recorded with no further additions (-), with added 1 mM KCN (o), 10,uM HQNO (A), or 5 mM sodium azide (o), or in equilibrium with an atmosphere of 0.2% 02 in N2 (o). Also shown is the effect the secondary addition of 1 mM cyanide (X) and 10 MM HQNO (A) during the luminescence decay. Luminescence is in light units; one unit = 1.7 X 108 quanta .isec-1.
Proc. Natl. Acad. Sci. USA 76 (1979)
Biochemistry: Ulitzur and Hastings The stimulatory effect of cyanide is not confined to the 14-carbon aldehyde. With the M17 mutant there is a large increase in the quantum yield of myristic acid in the presence of cyanide (Fig. 1), much more than with acids of other chain lengths (Table 1). The absolute photon yield with added tetradecanoic acid was always higher than with tetradecanal; quantum yields between 0.5 and 1 were commonly obtained with the acid when respiration was blocked. The reasons for this better yield may include solubility factors and the occurrence of so many competing side reactions for the aldehyde (13). With both acid and aldehyde (14 carbons) a similar stimulatory effect upon photon yields with mutant M17 occurs with other inhibitors of electron transport (HQNO and sodium azide) as well as under low (t0.2%) 02 (Fig. 2). These results indicate that the natural endogenous aldehyde in bacterial bioluminescence is the 14-carbon, saturated, aliphatic compound, and that there are cellular enzymes able to recycle its oxidation product (but not other chain lengths) by reduction. Although the results presented are for B. harveyi, similar observations in vivo indicate that the conclusions are also applicable for Photobacterium fischeri (8) and P. leiognathi. The small quantities of aldehyde that could be isolated from two species of luminous bacteria were mostly of chain lengths 12, 14, and 16 carbons, in relative abundance of 5, 63, and 30%, respectively, in one species and 36, 32, and 20% in the other (14). It is also interesting that with luciferase isolated from all three Photobacterium species the rate of the reaction is much greater with tetradecanal than with aldehydes of other
267
chain lengths, especially the naturally occurring even-numbered ones (ref. 15; unpublished data). 1. Strehler, B. L. & Cormier, J. J. (1953) Arch. Biochem. Biophys.
47,916-3.
2. Hastings, J. W. (1978) Photochem. Photobiol. 27, 397-404. 3. Hastings, J. W. & Nealson, K. H. (1977) Annu. Rev. Microbiol.
31,549-595. 4. Hastings, J. W., Weber, K., Friedland, J., Eberhard, A., Mitchell, F. W. & Gunsalus, A. (1969) Biochemistry 8, 4681-4689. 5. Rogers, P. & McElroy, W. D. (1955) Proc. Natl. Acad. Sci. USA 41, 67-70. 6. Cline, T. W. & Hastings, J. W. (1971) Proc. Natl. Acad. Sci. USA 68, 500-504. 7. Rogers, P. & McElroy, W. D. (1958) Arch. Biochem. Biophys.
75,87-105.
8. Ulitzur, S. & Hastings, J. W. (1978) Proc. Natl. Acad. Sci. USA
75,266-269.
9. Reichelt, J. L. & Baumann, P. (1973) Arch. Mikrobiol. 94,
283-330.
10. Mitchell, G. & Hastings, J. W. (1971) Anal. Biochem. 39, 243250. 11. Hastings, J. W. & Weber, G. (1963) J. Opt. Soc. Am. 53, 1410-1415. 12. Ulitzur, S. & Hastings, J. W. (1978) J. Bacteriol. 133, 13071313. 13. Schauenstein, H., Esterbauer, H. & Zollner, H. (1977) Aldehydes in Biological Systems (Pion, London). 14. Shimomura, O., Johnson, F. H. & Morise, H. (1974) Proc. NatI. Acad. Sci. USA 71, 4666-4669. 15. Hastings, J. W., Spudich, J. A. & Malnic, G. (1963) J. Biol. Chem.
a238,3100-3105.
