Biosynthesis Of Cytochromes Research Articles

Bacterial TANGO2 homologs are heme-trafficking proteins that facilitate biosynthesis of cytochromes c.

Heme, an essential molecule for virtually all living organisms, acts primarily as a cofactor in a large number of proteins. However, how heme is mobilized from the site of synthesis to the locations where hemoproteins are assembled remains largely unknown in cells, especially bacterial ones. In this study, with Shewanella oneidensis as the model, we identified HtpA (SO0126) as a heme-trafficking protein and homolog of TANGO2 proteins found in eukaryotes. We showed that HtpA homologs are widely distributed in all domains of living organisms and have undergone parallel evolution. In its absence, the cytochrome (cyt) c content and catalase activity decreased significantly. We further showed that both HtpA and representative TANGO2 proteins bind heme with 1:1 stoichiometry and a relatively low dissociation constant. Protein interaction analyses substantiated that HtpA directly interacts with the cytochrome c maturation system. Our findings shed light on cross-membrane transport of heme in bacteria and extend the understanding of TANGO2 proteins. IMPORTANCE The intracellular trafficking of heme, an essential cofactor for hemoproteins, remains underexplored even in eukaryotes, let alone bacteria. Here we developed a high-throughput method by which HtpA, a homolog of eukaryotic TANGO2 proteins, was identified to be a heme-binding protein that enhances cytochrome c biosynthesis and catalase activity in Shewanella oneidensis. HtpA interacts with the cytochrome c biosynthesis system directly, supporting that this protein, like TANGO2, functions in intracellular heme trafficking. HtpA homologs are widely distributed, but a large majority of them were found to be non-exchangeable, likely a result of parallel evolution. By substantiating the heme-trafficking nature of HtpA and its eukaryotic homologs, our findings provide general insight into the heme-trafficking process and highlight the functional conservation along evolution in all living organisms.

Differential gene content and gene expression for bacterial evolution and speciation of Shewanella in terms of biosynthesis of heme and heme-requiring proteins

BackgroundMost species of Shewanella harbor two ferrochelatase paralogues for the biosynthesis of c-type cytochromes, which are crucial for their respiratory versatility. In our previous study of the Shewanella loihica PV-4 strain, we found that the disruption of hemH1 but not hemH2 resulted in a significant accumulation of extracellular protoporphyrin IX (PPIX), but it is different in Shewanella oneidensis MR-1. Hence, the function and transcriptional regulation of two ferrochelatase genes, hemH1 and hemH2, are investigated in S. oneidensis MR-1.ResultIn the present study, deletion of either hemH1 or hemH2 in S. oneidensis MR-1 did not lead to overproduction of extracellular protoporphyrin IX (PPIX) as previously described in the hemH1 mutants of S. loihica PV-4. Moreover, supplement of exogenous hemins made it possible to generate the hemH1 and hemH2 double mutant in MR-1, but not in PV-4. Under aerobic condition, exogenous hemins were required for the growth of MR-1ΔhemH1ΔhemH2, which also overproduced extracellular PPIX. These results suggest that heme is essential for aerobic growth of Shewanella species and MR-1 could also uptake hemin for biosynthesis of essential cytochrome(s) and respiration. Besides, the exogenous hemin mediated CymA cytochrome maturation and the cellular KatB catalase activity. Both hemH paralogues were transcribed in wild-type MR-1, and the hemH2 transcription was remarkably up-regulated in MR-1ΔhemH1 mutant to compensate for the loss of hemH1. The periplasmic glutathione peroxidase gene pgpD, located in the same operon with hemH2, and a large gene cluster coding for iron, heme (hemin) uptake systems are absent in the PV-4 genome.ConclusionOur results indicate that the genetic divergence in gene content and gene expression between these Shewanella species, accounting for the phenotypic difference described here, might be due to their speciation and adaptation to the specific habitats (iron-rich deep-sea vent versus iron-poor freshwater) in which they evolved and the generated mutants could potentially be utilized for commercial production of PPIX.

Exogenous application of 5-aminolevulinic acid on wheat seedlings under drought stress enhances the transcription of psbA and psbD genes and improves photosynthesis

The compound 5-aminolevulinic acid (ALA) is an essential precursor for the biosynthesis of porphyrins, including chlorophyll, heme, and cytochromes. The protective effects of ALA on photosynthesis and the expression of photosynthetic genes in wheat under drought stress are not well understood. Two wheat cultivars, drought-tolerant Aikang-58 and drought-sensitive Chinese Spring, were exposed to drought stress induced by 20% polyethylene glycol (PEG-6000) after foliar pretreatment with ALA for 3 days. The results showed that exogenous application of ALA protected the drought-stressed wheat seedlings by significantly inhibiting the decrease in relative water and chlorophyll contents. The ALA-mediated alleviation of stress was similar between the drought-tolerant and drought-sensitive wheat cultivars. Meanwhile, compared to seedlings under drought treatment alone, the ALA-pre-treated wheat seedlings under drought stress maintained higher photosystem II (PSII) functional indexes. The ALA pretreatment reduced the drought-driven accumulation of both H2O2 and ABA and increased the stomatal conductance. Real-time PCR analysis showed that the psbA and psbD gene transcripts were both upregulated under drought stress following ALA pretreatment. The present study suggests that the exogenous foliar application of ALA alleviates the drought stress on wheat seedlings, which is associated with the enhancement of PSII function by inducing chlorophyll synthesis and psbA and psbD transcription. Moreover, the protective effect of ALA pretreatment was not related to the decline in stomatal conductance caused by ABA or H2O2 accumulation.

XC_0531 encodes a c-type cytochrome biogenesis protein and is required for pathogenesis in Xanthomonas campestris pv. campestris

BackgroundThe phytopathogenic Xanthomonas campestris pv.campestris is a gram-negative bacterium and the causal agent of black-rot disease of cruciferous crops. Many gram-negative bacteria possess a family of proteins, called Dsbs, which are involved in disulfide bond formation in certain periplasmic proteins. In our preliminary screening of the virulence to the plants we identified that gene XC_0531 which annotated gene dsbD of Xanthomonas campestris pv. campestris (Xcc) is related to the virulence to the host plants.ResultsHere, we found XC_0531 encoded a DsbD like protein. Its deletion is sensitive to DTT and copper, decreased accumulation of free thiols in periplasm. Its deletion also affected heme synthesis, position of Soret band and the production of peak c550. This suggests that XC_0531 is related to c-type cytochromes biogenesis. XC_0531 mutation decreased the utilization of different carbon sources (such as galactose, xylose, maltose, saccharose and glucose), reduced extracellular polysaccharide (EPS) production, decreased extracellular enzyme activities (protease, cellulose and amylase), slowed down growth rate of Xcc and weakened virulence to the plants. These results suggest that these phenotypes caused by XC_0531 mutation is possibly due to deficient biosynthesis of c-type cytochromes in respiration chain and the formation of disulfide bonds. Our work confirmed the function of XC_0531 and provide theory basis for scientists working on molecular mechanisms of cytochrome c biogenesis, pathogenesis of Xcc, development of EPS commercial values and protecting plant from black rot.ConclusionWe confirmed the function of gene XC_0531, which encodes a DsbD like protein, a protein correlated with c-type cytochrome biogenesis. This gene is related to the virulence to plants by affecting funtion of cytochromes c and probably disulfide bonds modification of proteins in type II secretion system (T2SS).

Biosynthesis of Single Thioether c-Type Cytochromes Provides Insight into Mechanisms Intrinsic to Holocytochrome c Synthase (HCCS).

C-type cytochromes (cyts c) are generally characterized by the presence of two thioether attachments between heme and two cysteine residues within a highly conserved CXXCH motif. Most eukaryotes use the System III cyt c biogenesis pathway composed of holocytochrome c synthase (HCCS) to catalyze thioether formation. Some protozoan organisms express a functionally equivalent, natural variant of cyt c with an XXXCH heme-attachment motif, resulting in a single covalent attachment. Previous studies have shown that recombinant HCCS can produce low levels of the XXXCH single thioether variant. However, cyt c variants containing substitutions at the C-terminal cysteine of the heme-attachment site (i.e., resulting in CXXXH) have never been observed in nature, and attempts to biosynthesize a recombinant version of this cyt c variant have been largely unsuccessful. In this study, we report the biochemical analyses of an HCCS-matured CXXXH cyt c variant, comparing its biosynthesis and properties to those of the XXXCH variant. The results indicate that although HCCS mediates heme attachment to the N-terminal cysteine in CXXXH cyt c variants, up to 50% of the cyt c produced is modified in an oxygen-dependent manner, resulting in a mixed population of cyt c. Since this aerobic modification occurs only in the context of CXXXH, we also propose that natural HCCS-mediated heme attachment to CXXCH likely initiates at the C-terminal cysteine.

Differential Regulation of the Two Ferrochelatase Paralogues in Shewanella loihica PV-4 in Response to Environmental Stresses.

Determining the function and regulation of paralogues is important in understanding microbial functional genomics and environmental adaptation. Heme homeostasis is crucial for the survival of environmental microorganisms. Most Shewanella species encode two paralogues of ferrochelatase, the terminal enzyme in the heme biosynthesis pathway. The function and transcriptional regulation of two ferrochelatase genes, hemH1 and hemH2, were investigated in Shewanella loihica PV-4. The disruption of hemH1 but not hemH2 resulted in a significant accumulation of extracellular protoporphyrin IX (PPIX), the precursor to heme, and decreased intracellular heme levels. hemH1 was constitutively expressed, and the expression of hemH2 increased when hemH1 was disrupted. The transcription of hemH1 was regulated by the housekeeping sigma factor RpoD and potentially regulated by OxyR, while hemH2 appeared to be regulated by the oxidative stress-associated sigma factor RpoE2. When an oxidative stress condition was mimicked by adding H2O2 to the medium or exposing the culture to light, PPIX accumulation was suppressed in the ΔhemH1 mutant. Consistently, transcriptome analysis indicated enhanced iron uptake and suppressed heme synthesis in the ΔhemH1 mutant. These data indicate that the two paralogues are functional in the heme synthesis pathway but regulated by environmental conditions, providing insights into the understanding of bacterial response to environmental stresses and a great potential to commercially produce porphyrin compounds. Shewanella is capable of utilizing a variety of electron acceptors for anaerobic respiration because of the existence of multiple c-type cytochromes in which heme is an essential component. The cytochrome-mediated electron transfer across cellular membranes could potentially be used for biotechnological purposes, such as electricity generation in microbial fuel cells and dye decolorization. However, the mechanism underlying the regulation of biosynthesis of heme and cytochromes is poorly understood. Our study has demonstrated that two ferrochelatase genes involved in heme biosynthesis are differentially regulated in response to environmental stresses, including light and reactive oxygen species. This is an excellent example showing how bacteria have evolved to maintain cellular heme homeostasis. More interestingly, the high yields of extracellular protoporphyrin IX by the Shewanella loihica PV-4 mutants could be utilized for commercial production of this valuable chemical via bacterial fermentation.

Zinc Deficiency Impacts CO2 Assimilation and Disrupts Copper Homeostasis in Chlamydomonas reinhardtii

Zinc is an essential nutrient because of its role in catalysis and in protein stabilization, but excess zinc is deleterious. We distinguished four nutritional zinc states in the alga Chlamydomonas reinhardtii: toxic, replete, deficient, and limited. Growth is inhibited in zinc-limited and zinc-toxic cells relative to zinc-replete cells, whereas zinc deficiency is visually asymptomatic but distinguished by the accumulation of transcripts encoding ZIP family transporters. To identify targets of zinc deficiency and mechanisms of zinc acclimation, we used RNA-seq to probe zinc nutrition-responsive changes in gene expression. We identified genes encoding zinc-handling components, including ZIP family transporters and candidate chaperones. Additionally, we noted an impact on two other regulatory pathways, the carbon-concentrating mechanism (CCM) and the nutritional copper regulon. Targets of transcription factor Ccm1 and various CAH genes are up-regulated in zinc deficiency, probably due to reduced carbonic anhydrase activity, validated by quantitative proteomics and immunoblot analysis of Cah1, Cah3, and Cah4. Chlamydomonas is therefore not able to grow photoautotrophically in zinc-limiting conditions, but supplementation with 1% CO2 restores growth to wild-type rates, suggesting that the inability to maintain CCM is a major consequence of zinc limitation. The Crr1 regulon responds to copper limitation and is turned on in zinc deficiency, and Crr1 is required for growth in zinc-limiting conditions. Zinc-deficient cells are functionally copper-deficient, although they hyperaccumulate copper up to 50-fold over normal levels. We suggest that zinc-deficient cells sequester copper in a biounavailable form, perhaps to prevent mismetallation of critical zinc sites.

Transmembrane heme delivery systems.

Heme proteins play pivotal roles in a wealth of biological processes. Despite this, the molecular mechanisms by which heme traverses bilayer membranes for use in biosynthetic reactions are unknown. The biosynthesis of c-type cytochromes requires that heme is transported to the bacterial periplasm or mitochondrial intermembrane space where it is covalently ligated to two reduced cysteinyl residues of the apocytochrome. Results herein suggest that a family of integral membrane proteins in prokaryotes, protozoans, and plants act as transmembrane heme delivery systems for the biogenesis of c-type cytochromes. The complete topology of a representative from each of the three subfamilies was experimentally determined. Key histidinyl residues and a conserved tryptophan-rich region (designated the WWD domain) are positioned at the site of cytochrome c assembly for all three subfamilies. These histidinyl residues were shown to be essential for function in one of the subfamilies, an ABC transporter encoded by helABCD. We believe that a directed heme delivery pathway is vital for the synthesis of cytochromes c, whereby heme iron is protected from oxidation via ligation to histidinyl residues within the delivery proteins.

The Plastid-encoded ccsA Gene Is Required for Heme Attachment to Chloroplast c-type Cytochromes

A chloroplast gene, ycf5, which displays limited sequence identity to bacterial genes (ccl1/cycK) required for the biogenesis of c-type cytochromes, was tested for its function in chloroplast cytochrome biogenesis in Chlamydomonas reinhardtii. Targeted inactivation of the ycf5 gene results in a non-photosynthetic phenotype attributable to the absence of c-type cytochromes. The cloned ycf5 gene also complements the phototrophic growth deficiency in strain B6 of C. reinhardtii. B6 is unable to synthesize functional forms of cytochromes f and c6 owing to a chloroplast genome mutation that prevents heme attachment. The selected (phototrophic growth) as well as the unselected (holocytochrome c6 accumulation) phenotypes were restored in complemented strains. The complementing gene, renamed ccsA (for c-type cytochrome synthesis), is expressed in wild-type and B6 cells but is non-functional in B6 owing to an early frameshift mutation. Antibodies raised against the ccsA gene product recognize a 29-kDa protein in C. reinhardtii. The 29-kDa protein is absent in strain B6 but is restored in a spontaneous revertant (B6R) isolated from a culture of B6. Sequence analysis of the ccsA gene in strain B6R indicates that it is a true revertant. We conclude that the ccsA gene is expressed and that it encodes a protein required for heme attachment to c-type cytochromes.

The biosynthesis of membrane and soluble plastidic c-type cytochromes of Chlamydomonas reinhardtii is dependent on multiple common gene products.

Cytochrome c6 functions in the thylakoid lumen to catalyze electron transfer from reduced cytochrome f of the cytochrome b6f complex to P700+ of photosystem I. The biogenesis of mature cyt c6 from cytosolically translated pre-apocytochrome c6 involves numerous post-translational modifications including the proteolytic removal of a transit sequence and the covalent attachment of heme to two cysteinyl thiols on the apoprotein. Here, we report on the characterization of a previously unrecognized class of non-allelic mutants of Chlamydomonas reinhardtii that are blocked at the conversion of apocyt c6 to holocyt c6. The mutants are acetate requiring since they are also deficient in cyt f, cyt b and the Rieske FeS protein. Pulse-chase studies indicate that heme attachment is not required for the two-step processing of pre-apocytochrome c6 to apocyt c6, but is required for the stability of the mature protein. This is in contrast to the biosynthesis of mitochondrial cyt c1 where heme attachment is required for the second processing step. We propose that the assembly of both holocytochrome c6 and the cytochrome b6f complex are dependent on common gene products, possibly those involved in heme delivery or metabolism. This is the first suggestion that multiple loci are involved in the biosynthesis of both plastidic c-type cytochromes.

The functions and biosynthesis of the c-type cytochromes of bacterial respiratory chains with particular reference to autotrophs and the handling of C 1 compounds

The behaviour, functions and biosynthesis of c-type cytochromes of the bacterial periplasm are reviewed. Particular emphasis is placed on cytochromes involved in (i) the metabolism of C 1 compounds and (ii) the electron transport reactions of Paracoccus denitrificans.

The functions and biosynthesis of thec-type cytochromes of bacterial respiratory chains with particular reference to autotrophs and the handling of C1compounds

The behaviour, functions and biosynthesis of c-type cytochromes of the bacterial periplasm are reviewed. Particular emphasis is placed on cytochromes involved in (i) the metabolism of C1 compounds and (ii) the electron transport reactions of Paracoccus denitrificans.

Apo forms of cytochrome C550 and cytochrome cd1 are translocated to the periplasm of Paracoccus denitrificans in the absence of haem incorporation caused by either mutation or inhibition of haem synthesis

An apo form of cytochrome c550 can be detected by immunoblotting cell-free extracts of a mutant of Paracoccus denitrificans that is deficient in c-type cytochromes. This apoprotein is found predominantly in the periplasm, the location of the holocytochrome in the wild-type organism, indicating that translocation of the polypeptide occurs in the absence of haem attachment. The polypeptide molecular weight, as judged by sodium dodecyl sulphate/polyacrylamide gel electrophoresis, is indistinguishable from that of the holoprotein and the chemically prepared apoprotein; this suggests that the N-terminal signal sequence is removed in the mutant as in the wild-type organism. In the presence of levulinic acid, an inhibitor of haem biosynthesis, apocytochrome c550 and aponitrite reductase (cytochrome cd1) accumulated in the periplasm of wild-type cells. Synthesis of these apoproteins was blocked by chloramphenicol. Thus in P. denitrificans the synthesis of these polypeptides is neither autoregulated nor regulated by the availability of haem. That the apoproteins appear in the periplasm argues against the possibility of polypeptide/haem co-transport from cytoplasm to periplasm. These observations are related to, and contrasted with, the biosynthesis of c-type cytochromes in eukaryotic cells.

Rhodobacter capsulatus MT113: A single mutation results in the absence of c-type cytochromes and in the absence of the cytochrome bc1 complex

MT113, a nonphotosynthetic mutant of Rhodobacter capsulatus previously characterized as lacking cytochrome c 2 is shown to lack also cytochrome c 1, the Rieske iron-sulfur cluster and the antimycin sensitive semiquinone Q ⨪ c, all components of the cytochrome bc 1 complex. Although MT113 contained b-type cytochromes and other iron-sulfur clusters at nearly wild-type level, it lacks c-type cytochromes. Based on antibody detection, c 2 apoprotein was absent in MT113, however the apoproteins corresponding to the cytochromes b and c 1 and the Rieske iron-sulfur cluster were present in reduced amounts. Genetic analysis indicated that the lesion appears to be due to a single mutation which is not localized in the structural genes of cytochrome c 2 or the bc 1 complex. These data taken together suggest that the pleiotropic mutation in MT113 might be related to the biosynthesis of c-type cytochromes.

Oxidative drug metabolism and inducibility by phenobarbital in sickle cell anemia.

The plasma elimination rates of antipyrine and phenylbutazone were studied in 8 patients with sickle cell anemia. The mean antipyrine half‐life (t½) in plasma was not significantly different in the sickle cell patients ( 12.0 ± 3.0 hr) than in a control group of 20 normal subjects (12.7 ± 0.9 hr). For phenylbutazone the mean tV2 was shorter in the sickle cell patients (2.1 ± 0.1 days) than in normal subjects (3.3 ± 0.2 days, p < 0.001); both mean apparent volume of distribution (aVD) and mean metabolic clearance rate (MCR) for this drug were significantly higher in the sickle cell group, which suggests increased extravascular distribution as well as more rapid hepatic metabolism. After administration of phenobarbital (2 to 3 mg/kg daily for 2 wk) consistent increases in rates of antipyrine and phenylbutazone metabolism were observed in 6 sickle cell anemia patients, which were comparable to those observed in 5 normal subjects. Two other patients who were taking drugs (methyltestosterone, isoniazid) known to alter cytochrome P‐450 function did not respond typically. Our results suggest that cytochrome P‐450 function and sensitivity to phenobarbital induction are unimpaired in most sickle cell anemia patients. Since uptake of heme into hepatocytes is probably increased in this disorder, this suggests that an adapative response develops during long‐term hemolysis such that heme biosynthesis and formation of microsomal cytochromes are not repressed.

Rapid and Economical Production of Microsomal Cytochrome P-450 in Yeast Resuspended in 20% Glucose Medium: Relationship to the Biosynthesis of Mitochondrial Cytochromes

Rapid and Economical Production of Microsomal Cytochrome <i>P</i>-450 in Yeast Resuspended in 20% Glucose Medium: Relationship to the Biosynthesis of Mitochondrial Cytochromes

Stimulation of fermentation and yeast-like morphogenesis in Mucor rouxii by phenethyl alcohol.

The germination of fungal spores into hyphae was inhibited by concentrations of phenethyl alcohol (PEA) from 0.05 to 0.3%. Spores of Mucor formed budding spherical cells instead of filaments. These cells were abundant in cultures of Mucor rouxii at 0.22% PEA, provided that the carbon source was a hexose at 2 to 5%. Morphology was filamentous with xylose, maltose, sucrose, or a mixture of amino acids. Removal of PEA resulted in the conversion of yeast-like cells into hyphae. PEA did not inhibit biosynthesis of cytochromes or oxygen uptake, but it stimulated CO(2) and ethyl alcohol production. PEA had no effect on the rate of oxygen uptake, but it inhibited the oxidative-phosphorylation activity of mitochondria. These results suggested that growth inhibition by PEA could result from uncoupling of oxidative phosphorylation and that, in Mucor, yeast-like morphology and fermentation were linked.

Altération par mutation de la régulation de la biosynthèse des cytochromes par les cultures anaérobies d'une bactérie anaérobie facultative

Starting with a wild strain of Hafnia having the phenotype: nitrate reductase A +, nitrate reductase B +, chlorate reductase C +, tetrathionate reductase +, fumarate reductase +, hydrogen lyase +, we have isolated two kinds of mutants having the following phenotypes: (1) A –, B +, C +, tetrathionate reductase +, fumarate reductase +, hydrogen lyase +; (2) A –, B –, C –, tetrathionate reductase –, fumarate reductase +, hydrogen lyase –. The wild strain and the mutants of category 1 can use nitrate, nitrite, fumarate, and tetrathionate as hydrogen acceptors under anaerobic conditions. The mutants of category 2 can also reduce nitrite and fumarate, but cannot use nitrate and tetrathionate as hydrogen acceptors.We have studied the effects of nitrate, nitrite, fumarate, and tetrathionate on the biosynthesis of cytochromes a2, b1, b‐562, and c‐550 using anaerobic cultures of the wild strain and several mutants of the above two categories. The following observations were made: (a) Wild strain. Neither tetrathionate nor fumarate have any effect. Nitrate, on the other hand, markedly reduces the levels of cytochromes a2, b‐562, and c‐550. As for nitrite, it causes a decrease in levels of cytochromes b‐562 and b1, but a strong increase in that of cytochrome c‐550. It is as though nitrate abolishes the effects of nitrite on the biosynthesis of cytochromes b1 and c‐550. (b) Mutants of category 1. Nitrate causes the cytochrome a2 band to disappear, but does not influence the biosynthesis of cytochromes b‐562 and c‐550. Apart from the effect of nitrate on the formation of cytochrome a2, the different hydrogen acceptors caused no detectable spectral changes. (c) Mutants of category 2. We obtain essentially the same spectra as with the wild strain except when the cultures are nitrite‐grown. In this case, nitrite has the same effect as nitrate because it represses the biosynthesis of cytochrome c‐550.It is clear from this work that the two mutant phenotypes studied have profound and different effects in their regulation of the biosynthesis of cytochromes.

CYTOCHROME PIGMENTS OF BAKER'S AND BREWER'S YEASTS

Baker's yeast, grown aerobically, has five absorption bands in the visible spectrum, due to cytochromes a and as at 605 mμ., cytochrome b at 565 and 530 mμ. and cytochrome c at 550 and 523 mμ. Brewer's yeast, anaerobically grown, can have up to three bands due to cytochrome a1 at approx. 590 mμ. and cytochrome b, at 558 and 530 mμ. Baker's yeast under anaerobic conditions becomes indistinguishable from brewer's yeast. Brewer's yeast grown aerobically becomes indistinguishable from baker's yeast and shows cytochromes a and a3, b and c. Cytochromes a, and b1 are said to be precursors in the biosynthesis of cytochromes a and a1, b and c A brewer's yeast incubated at 37° C. lost the cytochrome bands for a, and b1 and developed two new bands at 583 mμ. and 549 mμ. The compounds producing bands at 583 and 549 mμ. may be of biosynthetic importance because of their similarity to the spectra of compounds already reported in yeast, viz., haemoglobin and cytochrome c. Baker's yeast decreased in Qo2 value during fermentation but there was a rise in ability to produce carbon dioxide. The change in cytochrome constitution during anaerobiosis leaves a yeast with a low oxygen uptake. The ability to take up oxygen at all is probably only possible because of a reserve respiration system incorporating cytochromes a1 and b1.

Low-temperature spectral studies on the biosynthesis of cytochromes in baker's yeast

The time course of the biosynthesis of the cytochrome system was studied during aeration of anaerobically grown baker's yeast, spectrophotometrically at liquid-air temperature, and by measuring the rate of oxygen consumption with a polarized platinum electrode. Using cells harvested in the stationary phase of growth, the increase in Q o 2 values as well as the spectral transition from cytochrome b 1 to cytochromes b, C 1, and c was linear from the beginning of aeration, while in exponential phase cells a lag period of about an hour was observed. The lag was attributed in these cells to a lack of an oxygen-binding compound that serves as the inducer in the cytochrome-forming system. The low-temperature spectrum of reduced cytochrome b 1 in intact anaerobically grown yeast cells was shown to have two α bands, at 557.5 and 551.0 mμ, and to be similar to that of reduced cytochrome b 2, a purified enzyme from brewer's yeast, with bands of 557.5 and 552.5 mμ.