Kegg Pathway: Galactose metabolism

Environ Microbiol. 2009 Nov 3;
Schwalbach MS, Tripp HJ, Steindler L, Smith DP, Giovannoni SJ

Summary Bacteria in the SAR11 clade are highly abundant in marine surface waters, but currently little is known about the carbon compounds that support these large heterotrophic populations. To better understand the carbon requirements of these organisms, we conducted a multiphasic exploration of carbohydrate utilization among SAR11 isolates from the Northeast Pacific Ocean and the Sargasso Sea. A comparison of three SAR11 genomes showed they all lacked a recognizable PTS system, the oxidative portion of the pentose phosphate shunt (zwf(-), pgl(-)), genes for the Embden-Meyerhoff-Parnas (pfk(-), pyk(-)) and Entner-Doudoroff (eda(-)) pathways of glycolysis. Strain HTCC7211, isolated from an ocean gyre, was missing other glycolysis genes as well. Growth assays, radioisotopes, metagenomics and microarrays were used to test the hypothesis that these isolates might be limited in their abilities to transport and oxidize exogenous carbohydrates. Galactose, fucose, rhamnose, arabinose, ribose and mannose could not serve as carbon sources for the isolates tested. However, differences in glucose utilization were detected between coastal and ocean gyre isolates, with the coastal isolates capable of transporting, incorporating and oxidizing glucose while the open ocean isolate could not. Subsequent microarray analysis of a coastal isolate suggested that an operon encoding a variant of the Entner-Doudoroff pathway is likely responsible for the observed differences in glucose utilization. Metagenomic analysis indicated this operon is more commonly found in coastal environments and is positively correlated with chlorophyll a concentrations. Our results indicated that glycolysis is a variable metabolic property of SAR11 metabolism and suggest that glycolytic SAR11 are more common in productive marine environments.

BMC Mol Biol. 2009; 10: 99
Teste MA, Duquenne M, François JM, Parrou JL

BACKGROUND: Real-time RT-PCR is the recommended method for quantitative gene expression analysis. A compulsory step is the selection of good reference genes for normalization. A few genes often referred to as HouseKeeping Genes (HSK), such as ACT1, RDN18 or PDA1 are among the most commonly used, as their expression is assumed to remain unchanged over a wide range of conditions. Since this assumption is very unlikely, a geometric averaging of multiple, carefully selected internal control genes is now strongly recommended for normalization to avoid this problem of expression variation of single reference genes. The aim of this work was to search for a set of reference genes for reliable gene expression analysis in Saccharomyces cerevisiae. RESULTS: From public microarray datasets, we selected potential reference genes whose expression remained apparently invariable during long-term growth on glucose. Using the algorithm geNorm, ALG9, TAF10, TFC1 and UBC6 turned out to be genes whose expression remained stable, independent of the growth conditions and the strain backgrounds tested in this study. We then showed that the geometric averaging of any subset of three genes among the six most stable genes resulted in very similar normalized data, which contrasted with inconsistent results among various biological samples when the normalization was performed with ACT1. Normalization with multiple selected genes was therefore applied to transcriptional analysis of genes involved in glycogen metabolism. We determined an induction ratio of 100-fold for GPH1 and 20-fold for GSY2 between the exponential phase and the diauxic shift on glucose. There was no induction of these two genes at this transition phase on Galactose, although in both cases, the kinetics of glycogen accumulation was similar. In contrast, SGA1 expression was independent of the carbon source and increased by 3-fold in stationary phase. CONCLUSION: In this work, we provided a set of genes that are suitable reference genes for quantitative gene expression analysis by real-time RT-PCR in yeast biological samples covering a large panel of physiological states. In contrast, we invalidated and discourage the use of ACT1 as well as other commonly used reference genes (PDA1, TDH3, RDN18, etc) as internal controls for quantitative gene expression analysis in yeast.

Angew Chem Int Ed Engl. 2009; 48(47): 8848-69
Wennekes T, van den Berg RJ, Boot RG, van der Marel GA, Overkleeft HS, Aerts JM

The discovery of the glycosphingolipids is generally attributed to Johan L. W. Thudichum, who in 1884 published on the chemical composition of the brain. In his studies he isolated several compounds from ethanolic brain extracts which he coined cerebrosides. He subjected one of these, phrenosin (now known as galactosylceramide), to acid hydrolysis, and this produced three distinct components. One he identified as a fatty acid and another proved to be an isomer of D-glucose, which is now known as D-Galactose. The third component, with an "alkaloidal nature", presented "many enigmas" to Thudichum, and therefore he named it sphingosine, after the mythological riddle of the Sphinx. Today, sphingolipids and their glycosidated derivatives are the subjects of intense study aimed at elucidating their role in the structural integrity of the cell membrane, their participation in recognition and signaling events, and in particular their involvement in pathological processes that are at the basis of human disease (for example, sphingolipidoses and diabetes type 2). This Review details some of the recent findings on the biosynthesis, function, and degradation of glycosphingolipids in man, with a focus on the glycosphingolipid glucosylceramide. Special attention is paid to the clinical relevance of compounds directed at interfering with the factors responsible for glycosphingolipid metabolism.

IUBMB Life. 2009 Oct 26; 61(11): 1063-1074
Lai K, Elsas LJ, Wierenga KJ

In most organisms, productive utilization of Galactose requires the highly conserved Leloir pathway of Galactose metabolism. Yet, if this metabolic pathway is perturbed due to congenital deficiencies of the three associated enzymes, or an overwhelming presence of Galactose, this monosaccharide which is abundantly present in milk and many non-dairy foodstuffs, will become highly toxic to humans and animals. Despite more than four decades of intense research, little is known about the molecular mechanisms of Galactose toxicity in human patients and animal models. In this contemporary review, we take a unique approach to present an overview of Galactose toxicity resulting from the three known congenital disorders of Galactose metabolism and from experimental hyperGalactosemia. Additionally, we update the reader about research progress on animal models, as well as advances in clinical management and therapies of these disorders. (c) 2009 IUBMB IUBMB Life, 61(11): 1063-1074, 2009.

Med J Malaysia. 2009 Mar; 64(1): 83-5
Lee WS, Tay CG, Nazrul N, Paed M, Chai PF

A five-month-old Indian girl, product of consanguineous marriage, presented with diarrhoea with an onset within two days after birth, severe malnutrition and metabolic acidosis. The diarrhoea persisted even with lactose-free formula, amino acid-based formula and glucose-containing oral rehydration solution, but stopped when fasted. She required prolonged parenteral nutrition. Fructose and glucose tolerance tests were performed, confirming the child was able to absorb and metabolize fructose but not glucose, indicating a diagnosis of glucose-Galactose malabsorption. This case illustrate how simple and pertinent clinical observations and laboratory investigations is sufficient to allow a firm diagnosis to be made.

Microbiology. 2009 Oct 22;
Audy J, Labrie S, Roy D, Lapointe G

The effect of four sugars (glucose, Galactose, lactose, and fructose) on EPS production by B. longum subsp. longum CRC 002 was evaluated. More EPS was produced when CRC 002 was grown on lactose in the absence of pH control with a production of 1080 +/- 120 mg EPS l-1 (p < 0.01) after 24 h of incubation. For fructose, Galactose and glucose, EPS production was similar at 512 +/- 63 mg EPS l-1, 564 +/- 165 mg EPS l-1 and 612 +/- 93 mg EPS l-1, respectively. The proposed repeating unit composition of the EPS is 2 Galactose to 3 glucose. The effect of sugar and fermentation time on expression of genes involved in sugar nucleotide production (galK, galE1, galE2, galT1, galT2, galU, rmlA, rmlB1 and rmlCD) and the priming glycosyltransferase (wblE) was quantified using real time reverse transcription PCR (Q-RT-PCR). A significantly higher transcription level of wblE (9.29-fold) and the genes involved in the Leloir pathway (galK: 4.10-fold, galT1: 2.78-fold and galE2: 4.95-fold) during exponential growth are associated with enhanced EPS production on lactose compared to glucose. However, galU expression, linking glucose metabolism with the Leloir pathway, is not correlated with EPS production on different sugars. Genes coding for dTDP-rhamnose biosynthesis were also differentially expressed depending on sugar source and growth phase, although rhamnose was not present in the composition of the EPS. This precursor may be used in cell wall polysaccharide biosynthesis. These results contribute to understanding changes in gene expression when different sugar substrates are catabolized by B. longum subsp. longum CRC 002.

Infect Immun. 2009 Oct 19;
Terra VS, Homer KA, Rao SG, Andrew PW, Yesilkaya H

The pneumococcus obtains its energy from the metabolism of host glycosides. Therefore efficient degradation of host glycoproteins is integral to pneumococcal virulence. In search of novel pneumococcal glycosidases, we characterised the S. pneumoniae strain D39 protein encoded by SPD0065 and found that this gene encodes a beta-galactosidase. The SPD0065 recombinant protein released Galactose from desialylated fetuin, used here as a model of glycoproteins found in vivo. A pneumococcal mutant with a mutation in SPD0065 showed diminished beta-galactosidase activity, exhibited an extended lag period in mucin-containing defined medium and it cleaved significantly less Galactose than the parental strain during growth on mucin. As pneumococcal beta-galactosidase activity had been previously attributed solely to SPD0562 (bgaA), we evaluated the contribution of SPD0065 and SPD0562 to total beta-galactosidase activity. Mutation of either gene significantly reduced enzymic activity but beta-galactosidase activity in the double mutant, although significantly less than in either of the single mutants, was not completely abolished. The expression of SPD0065 in S. pneumoniae grown in mucin-containing medium or tissues harvested from infected animals was significantly up-regulated compared to pneumococci from glucose-containing medium. The SPD0065 mutant strain was found to be attenuated in virulence in a manner specific to the host tissue.

J Allergy Clin Immunol. 2009 Oct; 124(4): 652-7
Commins SP, Platts-Mills TA

Anaphylaxis is a severe allergic reaction that can rapidly progress and occasionally be fatal. In instances in which the triggering allergen is not obvious, establishing the cause of anaphylaxis is pivotal to long-term management. Assigning cause is limited, however, by the number of known exposures associated with anaphylaxis. Therefore identification of novel causative agents can provide an important step forward in facilitating new, allergen-specific approaches to management. In contrast to the view that carbohydrate-directed IgE has minimal, if any, clinical significance, recent data suggest that IgE antibodies to carbohydrate epitopes can be an important factor in anaphylaxis that might otherwise appear to be idiopathic. Here we review the evidence relating to carbohydrates in food allergy and anaphylaxis and discuss the implications of a new mammalian cross-reactive carbohydrate determinant.

Circ Cardiovasc Imaging. 2009 Jul; 2(4): 331-8
Laitinen I, Saraste A, Weidl E, Poethko T, Weber AW, Nekolla SG, Leppänen P, Ylä-Herttuala S, Hölzlwimmer G, Walch A, Esposito I, Wester HJ, Knuuti J, Schwaiger M

BACKGROUND: (18)F-Galacto-RGD is a positron emission tomography (PET) tracer binding to alpha(v)beta(3) integrin that is expressed by macrophages and endothelial cells in atherosclerotic lesions. Therefore, we evaluated (18)F-galacto-RGD for imaging vascular inflammation by studying its uptake into atherosclerotic lesions of hypercholesterolemic mice in comparison to deoxyglucose. METHODS AND RESULTS: Hypercholesterolemic LDLR(-/-)ApoB(100/100) mice on a Western diet and normally fed adult C57BL/6 control mice were injected with (18)F-galacto-RGD and (3)H-deoxyglucose followed by imaging with a small animal PET/CT scanner. The aorta was dissected 2 hours after tracer injection for biodistribution studies, autoradiography, and histology. Biodistribution of (18)F-galacto-RGD was higher in the atherosclerotic than in the normal aorta. Autoradiography demonstrated focal (18)F-galacto-RGD uptake in the atherosclerotic plaques when compared with the adjacent normal vessel wall or adventitia. Plaque-to-normal vessel wall ratios were comparable to those of deoxyglucose. Although angiogenesis was not detected, (18)F-galacto-RGD uptake was associated with macrophage density and deoxyglucose accumulation in the plaques. Binding to atherosclerotic lesions was efficiently blocked in competition experiments. In vivo imaging visualized (18)F-galacto-RGD uptake colocalizing with calcified lesions of the aortic arch as seen in CT angiography. CONCLUSIONS: (18)F-Galacto-RGD demonstrates specific uptake in atherosclerotic lesions of mouse aorta. In this model, its uptake was associated with macrophage density. (18)F-Galacto-RGD is a potential tracer for noninvasive imaging of inflammation in atherosclerotic lesions.

Zoolog Sci. 2009 Apr; 26(4): 249-53
Cao X, Mao D, Wang C, Zeng B, Wang A, Lu M, Xu C

A lectin with potent mitogenic activity was isolated from Musca domestica pupae. The lectin was a monomer with a molecular weight of 55 kDa. The purification procedure mainly involved affinity chromatography on Galactose-Sepharose-4B. The lectin agglutinated trypsin-treated rabbit blood cells. D-Galactose was detected to inhibit the lectin's hemagglutination activity. The lectin was metal independent and temperature dependent and was detected by the periodic acid-Shiff reaction to be a glycoprotein with a carbohydrate content of 2.1%. Observations with an atomic force microscope indicated that the lectin was a globular protein about 75 nm in size, with several carbohydrate chains linked to it. It elicited a mitogenic response from mouse splenocytes in vitro, with the maximal response at a concentration of 20 microg/ml. No lectin has been reported before from Musca domestica pupae, and the lectin we discovered could be of great importance in insect immunity and hypoimmune diseases.

J Proteome Res. 2009 Nov; 8(11): 4993-5007
Koskenniemi K, Koponen J, Kankainen M, Savijoki K, Tynkkynen S, de Vos WM, Kalkkinen N, Varmanen P

Lactobacillus rhamnosus GG (LGG) is one of the most extensively studied and widely used probiotic bacteria. While the benefits of LGG treatment in gastrointestinal disorders and immunomodulation are well-documented, functional genomics research of this bacterium has only recently been initiated. In the present study, a 2-D DIGE approach was used for the quantitative analysis of growth media-dependent changes in LGG protein abundance. Proteins were isolated from cells grown in industrial-type whey-based medium or in rich laboratory medium for subsequent 2-D DIGE. The analysis revealed patterns of protein abundance unique to each growth condition. In total, 196 quantitatively altered protein spots (at least 1.5-fold change in relative abundance, p < 0.05) representing approximately 13% of all protein spots in the gel were detected. From these protein spots, 157 were identified by mass spectrometry and were found to represent 100 distinct gene products. Collectively, these data show that growth of LGG in whey medium increased the relative abundance of proteins involved in purine biosynthesis, Galactose metabolism, and fatty acid biosynthesis. In comparison, growth of LGG in laboratory medium resulted in an increase in the amount of proteins involved in translation and the general stress response, as well as pyrimidine and exopolysaccharide biosynthesis. Moreover, several enzymes of the proteolytic system of LGG demonstrated growth medium-dependent production. The present study demonstrates the fundamental effects of culture conditions on the proteome of LGG, which are likely to affect the functionality and characteristics of its use as a probiotic.

Prikl Biokhim Mikrobiol. 2009 Jul-Aug; 45(4): 470-5
Giunter EA, Borisenkov MF, Ovodov IuS

Effects of UV-B (280-315 nm) and UV-C (254 nm) at various doses upon callus of bladder campion (Silene vulgaris (M.) G. were studied. It was revealed that UV irradiation results in the decrease in arabinose and Galactose residues in the silenan--the pectin fraction isolated from callus. The silenan possesses antioxidant activity (AOA) as assessed by the reaction with a stable radical. At the irradiation of callus by UV, the AOA of the silenan and the relative content of phenolic compounds in it increased; the highest increase was observed after the irradiation of callus by UV-B. Positive correlation between the AOA of the pectin fraction and an increase in phenolic compounds was revealed. This evidences that the AOA of the silenan relates to and is partially determined by phenolic compounds in its composition. The UV irradiation may be used as a tool to modify the structural features of the cell walls' polysaccharides in order to produce physiologically-active polysaccharides with desired properties.

Infect Immun. 2009 Dec; 77(12): 5418-27
Yesilkaya H, Spissu F, Carvalho SM, Terra VS, Homer KA, Benisty R, Porat N, Neves AR, Andrew PW

Knowledge of the in vivo physiology and metabolism of Streptococcus pneumoniae is limited, even though pneumococci rely on efficient acquisition and metabolism of the host nutrients for growth and survival. Because the nutrient-limited, hypoxic host tissues favor mixed-acid fermentation, we studied the role of the pneumococcal pyruvate formate lyase (PFL), a key enzyme in mixed-acid fermentation, which is activated posttranslationally by PFL-activating enzyme (PFL-AE). Mutations were introduced to two putative pfl genes, SPD0235 and SPD0420, and two putative pflA genes, SPD0229 and SPD1774. End-product analysis showed that there was no formate, the main end product of the reaction catalyzed by PFL, produced by mutants defective in SPD0420 and SPD1774, indicating that SPD0420 codes for PFL and SPD1774 for putative PFL-AE. Expression of SPD0420 was elevated in Galactose-containing medium in anaerobiosis compared to growth in glucose, and the mutation of SPD0420 resulted in the upregulation of fba and pyk, encoding, respectively, fructose 1,6-bisphosphate aldolase and pyruvate kinase, under the same conditions. In addition, an altered fatty acid composition was detected in SPD0420 and SPD1774 mutants. Mice infected intranasally with the SPD0420 and SPD1774 mutants survived significantly longer than the wild type-infected cohort, and bacteremia developed later in the mutant cohort than in the wild type-infected group. Furthermore, the numbers of CFU of the SPD0420 mutant were lower in the nasopharynx and the lungs after intranasal infection, and fewer numbers of mutant CFU than of wild-type CFU were recovered from blood specimens after intravenous infection. The results demonstrate that there is a direct link between pneumococcal fermentative metabolism and virulence.

Dev Biol. 2009 Nov 15; 335(2): 340-55
Novelli JF, Chaudhary K, Canovas J, Benner JS, Madinger CL, Kelly P, Hodgkin J, Carlow CK

Galactofuranose (Gal(f)), the furanoic form of d-Galactose produced by UDP-galactopyranose mutases (UGMs), is present in surface glycans of some prokaryotes and lower eukaryotes. Absence of the Gal(f) biosynthetic pathway in vertebrates and its importance in several pathogens make UGMs attractive drug targets. Since the existence of Gal(f) in nematodes has not been established, we investigated the role of the Caenorhabditis elegans UGM homolog glf-1 in worm development. glf-1 mutants display significant late embryonic and larval lethality, and other phenotypes indicative of defective surface coat synthesis, the glycan-rich outermost layer of the nematode cuticle. The glf homolog from the protozoan Leishmania major partially complements C. elegans glf-1. glf-1 mutants rescued by L. major glf, which behave as glf-1 hypomorphs, display resistance to infection by Microbacterium nematophilum, a pathogen of rhabditid nematodes thought to bind to surface coat glycans. To confirm the presence of Gal(f) in C. elegans, we analyzed C. elegans nucleotide sugar pools using online electrospray ionization-mass spectrometry (ESI-MS). UDP-Gal(f) was detected in wild-type animals while absent in glf-1 deletion mutants. Our data indicate that Gal(f) likely has a pivotal role in maintenance of surface integrity in nematodes, supporting investigation of UGM as a drug target in parasitic species.

Virol J. 2009; 6: 141
Yang B, Chuang H, Yang KD

BACKGROUND: Many viruses recognize specific sugar residues, particularly sulfated or sialylated glycans, as the infection receptors. A change of sialic acid (2-6)-linked Galactose (SA-alpha2,6Gal) to SA-alpha2,3Gal determines the receptor for avian flu infection. The receptor for enterovirus 71 (EV71) infection that frequently causes fatal encephalitis in Asian children remains unclear. Currently, there is no effective vaccine or anti-virus agent for EV71 infection. Using DLD-1 intestinal cells, this study investigated whether SA-linked glycan on DLD-1 intestinal cells was a receptor for EV71, and whether natural SA-linked sugars from human milk could block EV71 infection. RESULTS: EV71 specifically infected DLD-1 intestinal cells but not K562 myeloid cells. Depletion of O-linked glycans or glycolipids, but not N-linked glycans, significantly decreased EV71 infection of DLD-1 cells. Pretreatment of DLD-1 cells with sialidase (10 mU, 2 hours) significantly reduced 20-fold EV71 replication (p < 0.01). Taken together, these results suggest that SA-linked O-glycans and glycolipids, but not N-glycans, on DLD-1 cells were responsible for EV71 infection. Purified SA-alpha2,3Gal and SA-alpha2,6Gal from human milk significantly inhibited EV71 infection of DLD-1 cells, indicating terminal SA-linked glycans could be receptors and inhibitors of EV71 infection. CONCLUSION: This is the first in the literature to demonstrate that EV71 uses SA-linked glycans as receptors for infection, and natural SA-linked glycans from human milk can protect intestinal cells from EV71 infection. Further studies will test how a SA-containing glycan can prevent EV71 in the future.

Eur J Med Res. 2009 Sep 1; 14(9): 369-77
Miller M, Kahraman A, Ross B, Beste M, Gerken G

BACKGROUND AND AIMS: Quantitative tests of liver function (QTLF) which are based on the hepatic metabolism or clearance of test substances have been successfully used to predict prognosis of a variety of different liver diseases. Still sufficient data in HIV-patients under anti-retroviral therapy (ART) are lacking. Therefore, the aim of this prospective study was to investigate if and to what extent ART influences a broad panel of quantitative tests of liver function in patients with HIV-infection. PATIENTS AND METHODS: Nineteen patients (14 males, 5 females, mean age 40 years) with HIV-infection underwent QTLF including lidocaine half-life test (LHT), Galactose elimination capacity (GEC), and indocyanine green clearance (IGC). These tests were performed before and 3 to 6 months after initiation of anti-retroviral therapy. Twenty age-matched healthy, medication- and virus-free adults served as controls. RESULTS: Lidocaine half-life was significantly lower in HIV-patients without ART. Combining anti-retroviral therapies shifted cytochrome p450 activity back into standard ranges. Galactose elimination capacity as a parameter of cytosolic liver function and indocyanine green clearance as a parameter of liver perfusion were not affected by ART. CONCLUSIONS: QTLF may be a tool to predict prognosis or hepatic complications in HIV-infected patients with liver disease. Early determination of lidocaine half-life seems to be useful - this should be considered during the treatment of HIV-positive individuals.

Biochemistry (Mosc). 2009 Jul; 74(7): 709-16
Kawsar SM, Matsumoto R, Fujii Y, Yasumitsu H, Dogasaki C, Hosono M, Nitta K, Hamako J, Matsui T, Kojima N, Ozeki Y

A lectin was purified from Japanese sea hare Aplysia kurodai by lactosyl-agarose affinity chromatography. The molecular mass of the lectin was determined to be 56 and 32 kDa by SDS-PAGE under non-reducing and reducing conditions, respectively. It was found to agglutinate trypsinized and glutaraldehyde-fixed rabbit and human erythrocytes in the absence of divalent cations. The lectin exhibited stable thermo-tolerance as it retained hemagglutinating activity for 1 h even at 80 degrees C and showed stability at pH 10. By contrast, it was very sensitive at pH less than 5 and in the presence of the sulfhydryl-group preserving reagent, beta-mercaptoethanol. The hemagglutinating activity by the lectin was specifically inhibited by D-Galactose, galacturonic acid, methyl-alpha- and methyl-beta-D-galactopyranoside, lactose, melibiose, and asialofetuin. The association rate constant (k(ass)) and dissociation rate constant (k(diss)) were determined for the lectin to be 4.3 x 10(5) M(-1) x sec(-1) and 2.2 x 10(-3) sec(-1), respectively, using a surface plasmon resonance biosensor. The lectin moderately inhibited cell proliferation in the P388 cell line dose dependently. Interestingly, lectin-treated cells did not show a fragmented DNA ladder as is caused by apoptosis, suggesting that the cell proliferation inhibition was caused by another unknown mechanism.

Proteomics. 2009 Sep 9; 9(20): 4744-4754
Madinger CL, Sharma SS, Anton BP, Fields LG, Cushing ML, Canovas J, Taron CH, Benner JS

A proteomic analysis was performed on spent fermentation medium following bioreactor propagation of a wild-type industrial strain to identify proteins naturally secreted by Kluyveromyces lactis cells. Here, we report changes detected in the K. lactis secretome as a result of growth in three different carbon sources: glucose, Galactose and glycerol. A total of 151 secreted proteins were detected by multi-dimensional separations and reversed-phase online nanoESI-MS/MS analysis. From these, we were able to identify 63 proteins (termed the "base secretome") that were common to all three fermentation conditions. The majority of base secretome proteins, 79%, possessed general secretory pathway (GSP) sequences and were involved with cell wall structure, glycosylation, carbohydrate metabolism and proteolysis. There was little variation in the functional groupings of base secretome GSP proteins and GSP proteins that were not part of the base secretome. In contrast, the majority of non-GSP proteins detected were not part of the base secretome and the functions of these proteins varied significantly. Finally, through further identification of non-GSP proteins in carbon sources not originally tested, we have gained further evidence of a protein export mechanism separate from the GSP in K. lactis.

BMC Biotechnol. 2009; 9: 78
Oliveira C, Teixeira JA, Schmitt F, Domingues L

BACKGROUND: Numerous studies indicate that cancer cells present an aberrant glycosylation pattern that can be detected by lectin histochemistry. Lectins have shown the ability to recognise these modifications in several carcinomas, namely in the prostate carcinoma, one of the most lethal diseases in man. Thus, the aim of this work was to investigate if the alpha-D-Galactose-binding plant lectin frutalin is able to detect such changes in the referred carcinoma. Frutalin was obtained from different sources namely, its natural source (plant origin) and a recombinant source (Pichia expression system). Finally, the results obtained with the two lectins were compared and their potential use as prostate tumour biomarkers was discussed. RESULTS: The binding of recombinant and native frutalin to specific glycoconjugates expressed in human prostate tissues was assessed by using an immuhistochemical technique. A total of 20 cases of prostate carcinoma and 25 cases of benign prostate hyperplasia were studied. Lectins bound directly to the tissues and anti-frutalin polyclonal antibody was used as the bridge to react with the complex biotinilated anti-rabbit IgG plus streptavidin-conjugated peroxidase. DAB was used as visual indicator to specifically localise the binding of the lectins to the tissues. Both lectins bound to the cells cytoplasm of the prostate carcinoma glands. The binding intensity of native frutalin was stronger in the neoplasic cells than in hyperplasic cells; however no significant statistical correlation could be found (P = 0.051). On the other hand, recombinant frutalin bound exclusively to the neoplasic cells and a significant positive statistical correlation was obtained (P < 0.00001). However, recombinant frutalin did not recognise all malignant prostate cases and, when positive, the binding to those tissues was heterogeneous. CONCLUSION: Native and recombinant frutalin yielded different binding responses in the prostate tissues due to their differences in carbohydrate-binding affinities. Also, this study shows that both lectins may be used as histochemical biomarkers for the prostate cancer. Moreover, the successful use of a recombinant lectin in immunohistochemical studies of prostate cancer was for the first time demonstrated, highlighting the advantages of using recombinant systems in the preparation of pure lectin samples for diagnostic purpose.

J Allergy Clin Immunol. 2009 Sep; 124(3): 603-5
Jacquenet S, Moneret-Vautrin DA, Bihain BE