Maltase Gene Research Articles

Gene duplication and subsequent functional diversification of maltase in fig wasp (Chalcidoidea, Hymenoptera)

Maltase can catalyze the hydrolysis of α-1,4-glucosidic linkages and release α-d-glucoses that are used as a source of energy by insects. Maltase has been extensively studied in Lepidoptera and Diptera, while the characterization and evolutionary history of maltase are largely unknown in Hymenoptera. Here, we undertook a bioinformatics study and identified 105 maltase genes in 12 fig wasp species. Together with the maltase genes of Nasonia vitripennis and Apis mellifera, phylogenetic analysis showed that all the maltase genes were clustered into three clades. Clade I and III included maltase genes from all the fig wasp species, while clade II contained the maltase genes from non-pollinating fig wasps (NPFWs) only. Interestingly, the maltase genes located in clade II were intronless. Fig pollinators and NPFWs had lineage-specific gene expansion in clade I and II respectively, which were mainly derived from tandem duplications. The three clades displayed distinct gene structures. Furthermore, maltase showed significant functional divergence among the three clades and the critical amino acid sites were detected. These sites could be responsible for the ligand-binding preference and hydrolytic specificity. Overall, our results demonstrated that maltase might contribute to the discrepancy of life histories and feeding regimes between fig pollinators and NPFWs.

Sulfakinin inhibits activity of digestive enzymes in the brown planthopper, Nilaparvata lugens

• Two maltase genes and one lipase gene were identified in the brown planthopper. • Refed induces digestive enzyme secretion in Nilaparvata lugens . • Sulfakinin injection reduces digestive enzyme activity. • Sulfakinin injection decrease transcriptional levels of digestive enzyme genes. In animals, feeding can regulate release of digestive enzymes. Digestive enzymes are produced and released in response to specific ratios of nutrients, so the quality and quantity of food ingested are important factors in the secretion and activity of digestive enzymes. In general, the enzyme activity and secretion in the fed insects are relatively higher than that in the unfed insects. Neuropeptides and peptide hormones are important regulators of enzyme activity. In several insects, the neuropeptide sulfakinin (SK) is known to be a regulator of feeding and digestion similar to cholecystokinin in mammals. However, the roles of diet and SK in regulation of activity of digestive enzymes in the important pest insect, the brown planthopper ( Nilaparvata lugens ), are unknown. In this study, we identified six genes encoding different digestive enzymes and cloned three of these. We found that enzymatic activity and transcriptional levels of digestive enzymatic activity genes were upregulated by refeeding animals for 5 h after 24 h starvation. Furthermore, injection of N. lugens SK reduces digestive enzyme activity and leads to a downregulation of digestive enzyme gene transcripts. This study provides new views into the action of diet and SK in regulation of digestive enzymes in (hemimetabolous) insects. Taken together with the roles of SK in inducing satiety, our data strongly suggest that SK signaling is important in regulation of food ingestion and processing.

The ploidy determination of the biotechnologically important yeast Candida utilis.

Yeast Candida utilis is considered to be a potentially advantageous expression system for production of recombinant proteins utilizable for industrial and pharmaceutical purposes. As the scientific literature is not consistent in the ploidy of this yeast, in this work, we focused on resolving the problem via several methods such as the copy number determination of maltase gene by multiplex PCR, measuring α-glucosidase activity, the characterization of maltase gene copy number in deletion mutants using qPCR and flow cytometry. In context with the published data and results obtained in this study about the copy number of the maltase gene on C. utilis genome, we attempted to hypothesise and made conclusion about the ploidy of C. utilis. The results of this work, besides the biotechnological aspect, contribute to the elementary knowledge of C. utilis. The exact information about the ploidy or more specifically about the copy number of appropriate gene is essential for expression cassette dosage determination integrated into the chromosome of the host. In this study, we come to the conclusion that the maltase gene is present in C. utilis genome in four alleles, and in combination with flow cytometry, published information and the published genome sequences, the observations support the theory about tetraploidy of C. utilis.

Essentiality of the Maltase AmlE in Maltose Utilization and Its Transcriptional Regulation by the Repressor AmlR in the Acarbose-Producing Bacterium Actinoplanes sp. SE50/110.

Actinoplanes sp. SE50/110 is the wild type of industrial production strains of the fine-chemical acarbose (acarviosyl-maltose), which is used as α-glucosidase inhibitor in the treatment of type II diabetes. Although maltose is an important building block of acarbose, the maltose/maltodextrin metabolism has not been studied in Actinoplanes sp. SE50/110 yet. Bioinformatic analysis located a putative maltase gene amlE (ACSP50_2474, previously named malL; Wendler et al., 2015a), in an operon with an upstream PurR/LacI-type transcriptional regulator gene, named amlR (ACSP50_2475), and a gene downstream (ACSP50_2473) encoding a GGDEF-EAL-domain-containing protein putatively involved in c-di-GMP signaling. Targeted gene deletion mutants of amlE and amlR were constructed by use of the CRISPR/Cas9 technology. By growth experiments and functional assays of ΔamlE, we could show that AmlE is essential for the maltose utilization in Actinoplanes sp. SE50/110. Neither a gene encoding a maltose phosphorylase (MalP) nor MalP enzyme activity were detected in the wild type. By this, the maltose/maltodextrin system appears to be fundamentally different from other described prokaryotic systems. By sequence similarity analysis and functional assays from the species Streptomyces lividans TK23, S. coelicolor A3(2) and S. glaucescens GLA.O, first hints for a widespread lack of MalP and presence of AmlE in the class Actinobacteria were given. Transcription of the aml operon is significantly repressed in the wild type when growing on glucose and repression is absent in an ΔamlR deletion mutant. Although AmlR apparently is a local transcriptional regulator of the aml operon, the ΔamlR strain shows severe growth inhibitions on glucose and – concomitantly – differential transcription of several genes of various functional classes. We ascribe these effects to ACSP50_2473, which is localized downstream of amlE and presumably involved in the metabolism of the second messenger c-di-GMP. It can be assumed, that maltose does not only represent the most important carbon source of Actinoplanes sp. SE50/110, but that its metabolism is coupled to the nucleotide messenger system of c-di-GMP.

Association between duplicated maltase genes and the transcriptional regulation for the carbohydrate changes in Drosophila melanogaster

Gene duplication could promote phenotypic and genetic adaptation to various environments. To understand the effects of gene duplication on transcriptional regulation associated with environmental changes, we focused on the starch hydrolysis pathway, in which amylase enzymes together with maltase enzymes hydrolyze starch into glucose. Drosophila genomes involve ten duplicated Maltase genes. We examined the levels of transcription of the nine of these genes in 36 lines of Drosophila melanogaster collected from a natural population. In the investigated population, the levels of transcription were different between the two dietary carbohydrate sources, glucose and starch. At the transcriptional level, a single Maltase gene, which transcribes the specific Maltase transcripts, worked together with an Amylase gene in the pathway. The three of nine genes responded to carbohydrate changes, and the degree of the response was similar to Amylase gene. Our results suggest that gene duplication could increase capacity of the transcriptional regulation associated with environmental changes.

A novel sucrose-based expression system for heterologous proteins expression in thermotolerant methylotrophic yeast Ogataea thermomethanolica.

The thermotolerant methylotrophic yeast Ogataea thermomethanolica TBRC656 is a potential host for heterologous protein expression. In this study, a novel expression system was developed for O. thermomethanolica based on the maltase (mal) gene promoter from this organism. The OtMal promoter function was tested for expression of fungal enzymes as reporter genes. Measurement of xylanase reporter enzyme activity showed that the OtMal promoter was repressed during growth on glucose and was activated by sucrose. When sucrose was used as a carbon source, the OtMal promoter was approximately twice as strong as the constitutive OtGAP promoter. Comparison of the OtMal promoter with the methanol-inducible OtAOX promoter showed that OtMal promoter drove 1.2 and 1.7-fold higher expression of xylanase and phytase reporter, respectively, than OtAOX promoter under inducing conditions at 24 h. Our results indicated that this novel expression system could be useful for the production of heterologous proteins from sucrose in yeast O. thermomethanolica.

Molecular Regulations of Mucosal Maltase Expressions.

Two major α-glucosidase (maltase) genes, sucrase-isomaltase (SI) and maltase-glucoamylase (MGAM), respectively, are expressed in the small intestine. In this review, we have summarized whether jejunal expression of these maltase genes is regulated by dietary manipulations, which may affect carbohydrate availability from the luminal side, through changes in the binding of transcription factors and/or histone code on these genes. Studies using a model of mice fed either a low-starch or a high-starch diet for 7 days, found the mRNA levels of SI, MGAM, and Na-glucose cotransporter (SGLT1) genes in the jejunum to be increased in parallel by feeding a high-starch diet. Chromatin immunoprecipitation assays, using jejunal tissue of mice and rats fed a high-starch diet, revealed that the diet increased the acetylations of histones H3 and H4, bindings of coactivators, including general control of amino acid synthesis (GCN5) and the transcriptional factors, including caudal-related homeobox 2 (CDX2), and hepatocyte nuclear factor 1 (HNF1), not only in the promoter/enhancer regions, but also in the transcribed regions of SI and MGAM genes. Feeding rats a diet rich in resistant starch led to a concomitant reduction of mRNA levels of the MGAM gene and histone H3 modifications (acetylations and di-/tri-methylations) in the jejunum. These data suggest that a signal elicited by available glucose in the jejunal mucosa is associated with SI and MGAM gene expressions through a histone code, such as acetylation and di-/tri-methylations of histone H3 in the promoter/enhancer and transcribed regions of SI and MGAM genes.

Molecular machinery of starch digestion and glucose absorption along the midgut of Musca domestica

Until now there is no molecular model of starch digestion and absorption of the resulting glucose molecules along the larval midgut of Musca domestica. For addressing to this, we used RNA-seq analyses from seven sections of the midgut and carcass to evaluate the expression level of the genes coding for amylases, maltases and sugar transporters (SP). An amylase related protein (Amyrel) and two amylase sequences, one soluble and one with a predicted GPI-anchor, were identified. Three highly expressed maltase genes were correlated with biochemically characterized maltases: one soluble, other glycocalyx-associated, and another membrane-bound. SPs were checked as being apical or basal by proteomics of microvillar preparations and those up-regulated by starch were identified by real time PCR. From the 9 SP sequences with high expression in midgut, two are putative sugar sensors (MdSP4 and MdSP5), one is probably a trehalose transporter (MdSP8), whereas MdSP1-3, MdSP6, and MdSP9 are supposed to transport glucose into cells, and MdSP7 from cells to hemolymph. MdSP1, MdSP7, and MdSP9 are up-regulated by starch. Based on the data, starch is at first digested by amylase and maltases at anterior midgut, with the resulting glucose units absorbed at middle midgut. At this region, low pH, lysozyme, and cathepsin D open the ingested bacteria and fungi cells, freeing sugars and glycogen. This and the remaining dietary starch are digested by amylase and maltases at the end of middle midgut and up to the middle part of the posterior midgut, with resulting sugars being absorbed along the posterior midgut.

The Maltase Involved in Starch Metabolism in Barley Endosperm Is Encoded by a Single Gene.

During germination and early seedling growth of barley (Hordeum vulgare), maltase is responsible for the conversion of maltose produced by starch degradation in the endosperm to glucose for seedling growth. Despite the potential relevance of this enzyme for malting and the production of alcoholic beverages, neither the nature nor the role of maltase is fully understood. Although only one gene encoding maltase has been identified with certainty, there is evidence for the existence of other genes and for multiple forms of the enzyme. It has been proposed that maltase may be involved directly in starch granule degradation as well as in maltose hydrolysis. The aim of our work was to discover the nature of maltase in barley endosperm. We used ion exchange chromatography to fractionate maltase activity from endosperm of young seedlings, and we partially purified activity for protein identification. We compared maltase activity in wild-type barley and transgenic lines with reduced expression of the previously-characterised maltase gene Agl97, and we used genomic and transcriptomic information to search for further maltase genes. We show that all of the maltase activity in the barley endosperm can be accounted for by a single gene, Agl97. Multiple forms of the enzyme most likely arise from proteolysis and other post-translational modifications.

Upstream regulatory regions controlling the expression of the Candida utilis maltase gene

Candida utilis represents a promising expression host, generating relatively high levels of recombinant proteins. The current study presents preliminary characterization of the upstream regulatory regions controlling the carbon source-dependent expression of the C. utilis maltase gene. Cellobiose and soluble starch were recognised as inducers of maltase promoter, besides maltose. We successfully applied the Cre-loxP system to acquire a null mutant strain with disrupted maltase gene and promoter in polyploid yeast C. utilis. Furthermore, the strength and minimal functional region of the promoter was evaluated by measuring β-galactosidase activity. Our results suggest, the qPCR was shown itself a very smart method for relatively easy characterization of the transformants and correlation of the expression levels with gene dosage.

The effect of carbon sources on upstream regulatory regions controlling the expression of the Candida utilis maltase gene

The effect of carbon sources on upstream regulatory regions controlling the expression of the Candida utilis maltase gene

Characterization of Maltase Clusters in the Genus Drosophila

To reveal evolutionary history of maltase gene family in the genus Drosophila, we undertook a bioinformatics study of maltase genes from available genomes of 12 Drosophila species. Molecular evolution of a closely related glycoside hydrolase, the α-amylase, in Drosophila has been extensively studied for a long time. The α-amylases were even used as a model of evolution of multigene families. On the other hand, maltase, i.e., the α-glucosidase, got only scarce attention. In this study, we, therefore, investigated spatial organization of the maltase genes in Drosophila genomes, compared the amino acid sequences of the encoded enzymes and analyzed the intron/exon composition of orthologous genes. We found that the Drosophila maltases are more numerous than previously thought (ten instead of three genes) and are localized in two clusters on two chromosomes (2L and 2R). To elucidate the approximate time line of evolution of the clusters, we estimated the order and dated duplication of all the 10 genes. Both clusters are the result of ancient series of subsequent duplication events, which took place from 352 to 61million years ago, i.e., well before speciation to extant Drosophila species. Also observed was a remarkable intron/exon composition diversity of particular maltase genes of these clusters, probably a result of independent intron loss after duplication of intron-rich gene ancestor, which emerged well before speciation in a common ancestor of all extant Drosophila species.

Hsp90 Cochaperone Aha1 Is a Negative Regulator of the Saccharomyces MAL Activator and Acts Early in the Chaperone Activation Pathway

Aha1 is a ubiquitous cochaperone of the Hsp90/Hsp70 chaperone machine. It binds the middle domain of Hsp90 and stimulates ATPase activity, suggesting a function late in the chaperone pathway. Saccharomyces Mal63 MAL activator is a DNA-binding transcription factor and Hsp90 client protein. This study utilizes several MAL activator mutants to investigate Aha1 function in vivo. Deletion of AHA1 enhances induced Mal63-dependent maltase activity levels 2-fold, whereas overproduction of Aha1 represses expression. Maltase expression in strains carrying constitutive and super-inducible mutant activators with alterations near the C terminus (particularly residues 433-463) is unaffected by either aha1Delta or Aha1 overproduction. However, another constitutive activator with alterations outside of this C-terminal region is sensitive to Aha1 regulation. Previously, we showed that in the absence of inducer, Mal63 forms a stable intermediate complex with Hsp70, Hsp90, and Sti1, whereas noninducible mutant activators bind only with Hsp70 in an apparent early complex. Here, we find that triple Myc-tagged Aha1/Myc3 copurifies with all noninducible Mal63 mutant activators tested. Aha1/Myc3 association with inducible Mal63 is observed only in a sti1Delta strain, in which Hsp90 binding and intermediate complex formation are defective. Constitutive and super-inducible mutant activators with C-terminal alterations do not bind Aha1 even in a sti1Delta strain. Mal63 binding to Hsp90 and Hsp70 is enhanced 3-fold by loss of Aha1. These results suggest an interaction between Aha1 and residues near the C terminus of Mal63 thereby regulating Hsp90 association. A novel mechanism for the negative regulation of the MAL activator by Aha1 cochaperone is proposed.

Cloning and overexpression of a maltase gene from Schizosaccharomyces pombe in Escherichia coli and characterization of the recombinant maltase

The Schizosaccharomyces pombe maltase structural gene ( SPMAL1 +) was amplified from genomic DNA of S. pombe by PCR. An open reading frame of 1740 bp, encoding a putative 579 amino-acid protein with a calculated molecular mass of 67.7 kDa was characterized in the genomic DNA insert of plasmid pQE30. The specific maltase activity in the induced transformants was 21 times higher than that in wild-type. However, the estimated molecular mass of the purified recombinant maltase was 44.3 kDa by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The optimal temperature and pH of the purified recombinant maltase were 40 °C and 6, respectively. The recombinant maltase was weakly activated by Mg 2+, Ca 2+, Na +, and Ba 2+, but was strongly inhibited by Hg 2+, Ag + and Cu 2+, EDTA, and PMSF. The purified maltase could actively hydrolyse ρ-nitrophenyl glucoside (PNPG), maltose, dextrin, and soluble starch. The results demonstrate that maltase from S. pombe was different from that from other yeasts, and might be usefully exploited in the future by the biotechnology industry or lead to the development of new molecular genetic tools.

Hansenula polymorpha maltase gene promoter with sigma 70-like elements is feasible for Escherichia coli-based biotechnological applications: Expression of three genomic levansucrase genes of Pseudomonas syringae pv. tomato

P MAL1, the sigma 70-like sequences possessing promoter of the maltase gene of Hansenula polymorpha, was evaluated for its application in heterologous protein production in Escherichia coli. Levansucrases Lsc1, Lsc2 and Lsc3 of Pseudomonas syringae pv. tomato DC3000 were expressed from P MAL1 in E. coli as biotechnologically relevant model proteins. Production of soluble levansucrases with high specific activity confirmed appropriate strength of P MAL1 in a prokaryotic host. As about 90% of levansucrase activity was present in the cytoplasm of E. coli, no levan-synthesis related sucrose intolerance of bacteria was observed. All three levansucrases hydrolyzed and polymerized both, sucrose and raffinose. The raffinose-related activity of levansucrases has not been previously described in P. syringae. The P MAL1 expression system was used to produce Lsc3 protein in E. coli for purification. The purified levansucrase showed much higher affinity for sucrose cleavage ( K m = 21 mM) than the levansucrase from P. syringae pv. phaseolicola described in the literature with K m of 160 mM.

Studies on inositol-mediated expression of MAL gene encoding maltase and phospholipid biosynthesis in Schizosaccharomyces pombe

In this study, the effects of inositol addition on expression of the MAL gene encoding maltase and phosphatidylinositol (PI) biosynthesis in Schizosaccharomyces pombe (a naturally inositol-requiring strain) were examined. We found that specific maltase activity was at its maximum when the concentration of added inositol reached 6 microg ml(-1) in a synthetic medium containing 2.0% (w/v) glucose. When the concentration of added inositol was 1 microg ml(-1) in the medium, repression of MAL gene expression occurred at glucose concentration higher than 0.2% (w/v). However, when S. pombe was cultured in the synthetic medium containing 6 microg ml(-1), repression of maltase gene expression occurred only at initial glucose concentration above 1.0% (w/v). More mRNA encoding maltase was detected in the cells grown in the medium with 6 microg ml(-1) inositol than in those grown in the same medium with 1 microg ml(-1) inositol. These results demonstrate that higher inositol concentrations in the synthetic medium could derepress MAL gene expression in S. pombe. PI content of the yeast cells grown in the synthetic medium with 6 microg ml(-1) of inositol was higher than that of the yeast cells grown in the same medium with 1 microg ml(-1) of inositol. This means that PI may be involved in the derepression of MAL gene expression in S. pombe.

Clustering of genes in : Cloning of the maltose permease gene and expression from the divergent intergenic region between the maltose permease and maltase genes

Hansenula polymorpha uses maltase to grow on maltose and sucrose. Inspection of genomic clones of H. polymorpha showed that the maltase gene HPMAL1 is clustered with genes corresponding to Saccharomyces cerevisiae maltose permeases and MAL activator genes orthologues. We sequenced the H. polymorpha maltose permease gene HPMAL2 of the cluster. The protein (582 amino acids) deduced from the HPMAL2 gene is predicted to have eleven transmembrane domains and shows 39-57% identity with yeast maltose permeases. The identity of the protein is highest with maltose permeases of Debaryomyces hansenii and Candida albicans. Expression of the HPMAL2 in a S. cerevisiae maltose permease-negative mutant CMY1050 proved functionality of the permease protein encoded by the gene. HPMAL1 and HPMAL2 genes are divergently positioned similarly to maltase and maltose permease genes in many yeasts. A two-reporter assay of the expression from the HPMAL1-HPMAL2 intergenic region showed that expression of both genes is coordinately regulated, repressed by glucose, induced by maltose, and that basal expression is higher in the direction of the permease gene.

Regulation of the Hansenula polymorpha maltase gene promoter in H. polymorpha and Saccharomyces cerevisiae1.

Hansenula polymorpha is an exception among methylotrophic yeasts because it can grow on the disaccharides maltose and sucrose. We disrupted the maltase gene (HPMAL1) in H. polymorpha 201 using homologous recombination. Resulting disruptants HP201HPMAL1Delta failed to grow on maltose and sucrose, showing that maltase is essential for the growth of H. polymorpha on both disaccharides. Expression of HPMAL1 in HP201HPMAL1Delta from the truncated variants of the promoter enabled us to define the 5'-upstream region as sufficient for the induction of maltase by disaccharides and its repression by glucose. Expression of the Saccharomyces cerevisiae maltase gene MAL62 was induced by maltose and sucrose, and repressed by glucose if expressed in HP201HPMAL1Delta from its own promoter. Similarly, the HPMAL1 promoter was recognized and correctly regulated by the carbon source in a S. cerevisiae maltase-negative mutant 100-1B. Therefore we suggest that the transcriptional regulators of S. cerevisiae MAL genes (MAL activator and Mig1 repressor) can affect the expression of the H. polymorpha maltase gene, and that homologues of these proteins may exist in H. polymorpha. Using the HPMAL1 gene as a reporter in a H. polymorpha maltase disruption mutant it was shown that the strength of the HPMAL1 promoter if induced by sucrose is quite comparable to the strength of the H. polymorpha alcohol oxidase promoter under conditions of methanol induction, revealing the biotechnological potential of the HPMAL1 promoter.

Adeno-associated virus-mediated transfer of human acid maltase gene results in a transient reduction of glycogen accumulation in muscle of Japanese quail with acid maltase deficiency.

Glycogen storage disease type II (GSD II), Pompe's disease, is caused by the deficiency of acid alpha-D-glucosidase (GAA) in lysosome and is the most common form of GSD in Taiwan. Most cases are the infantile form. The disease is relentless and most patients die of cardiac failure and respiratory tract infection in the first year of life. At present, no treatment has been proved effective for this fatal disease. The applicability of enzyme replacement therapy is under investigation. However, high price and transient efficiency are the major problems to be solved. Accordingly, gene therapy by viral method has been conducted. In this study we constructed a plasmid that contained 5'-shortened BglII-NotI fragment human GAA cDNA, downstream of CMV promoter and bovine growth hormone polyadenylation signal, as well as AAV ITR region. When fibroblasts obtained from GSD II patients were cultured and infected with rAAV-GAA, the GAA activity of the fibroblasts increased four- to five-fold. Using acid maltase deficient (AMD) Japanese quail as the animal model, rcAAV-GAA 0.1 ml per site (1 x 10(9)-10) particles), totally 10 different sites to make 1 ml (1 x 10(1)0-11) particles), was injected into unilateral deep pectoral muscle of AMD quails. Medium (hepes) was only injected in the same way into the contralateral deep pectoral muscle to serve as control. Four days after injection, PAS staining showed disappearance of the glycogenosomes with regeneration of myocytes surrounding the intramuscular injected area as compared with the contralateral muscle of the same birds. Using anti-GAA monoclonal antibody, GAA was demonstrated on the regenerated myocytes by immunohistochemical staining and absent on the contralateral muscle of the same birds. Nevertheless, T lymphocytes infiltration was noted in both the rcAAV-GAA and hepes (medium) injected muscles and more prominent in the rcAAV-GAA-injected site. Functional evaluation demonstrated that wing flapping movement improved with wide flapping in the rAAV-GAA injected side, but not in the counterpart. Unfortunately, these histochemical and functional improvements faded away in 14 days, probably due to destruction of rcAAV by cell-mediated immunity of infiltrated T cells. Taken together, the present study suggests that rAAV can enter either human or quail cells and express and effectively reduce the glycogen accumulation in the skeletal muscle of AMD quails. These preliminary results are similar to these of low-dose rGAA replacement therapy. The mechanisms underlying the induction of cell-mediated immunity are unknown. How to elevate the number of packaged AAV, enhance the infectivity of AAV and reduce cell-mediated immunity must be solved in the future.

Respiration-Dependent Utilization of Sugars in Yeasts: a Determinant Role for Sugar Transporters

In many yeast species, including Kluyveromyces lactis, growth on certain sugars (such as galactose, raffinose, and maltose) occurs only under respiratory conditions. If respiration is blocked by inhibitors, mutation, or anaerobiosis, growth does not take place. This apparent dependence on respiration for the utilization of certain sugars has often been suspected to be associated with the mechanism of the sugar uptake step. We hypothesized that in many yeast species, the permease activities for these sugars are not sufficient to ensure the high substrate flow that is necessary for fermentative growth. By introducing additional sugar permease genes, we have obtained K. lactis strains that were capable of growing on galactose and raffinose in the absence of respiration. High dosages of both the permease and maltase genes were indeed necessary for K. lactis cells to grow on maltose in the absence of respiration. These results strongly suggest that the sugar uptake step is the major bottleneck in the fermentative assimilation of certain sugars in K. lactis and probably in many other yeasts.