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1.
Food Chem ; 301: 125271, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31376690

RESUMO

Mature endosperm was separated regionally into different parts in three rice cultivars, Te-qing (TQ), Wu-xiang 9915 (WX9915) and Guang-ling-xiang-nuo (GLXN), and their transgenic lines with inhibition of starch branching enzyme I and IIb (SBEI/IIb-). Within the three wild-type cultivars, starches from endosperm different regions showed similar molecular and crystalline structures. However, in rices with inhibition of SBEs, amylopectin short branch-chain content and branching degree gradually decreased, but amylopectin B3+ chain content and average chain length increased gradually from the interior to exterior of endosperm. The amylose content gradually increased from the interior to exterior of endosperm in TQ- and WX9915-SBEI/II- lines. From the interior to exterior of endosperm, starch changed gradually from CC- to CB-type in TQ-SBEI/II- line and from CA- to CC-type in GLXN-SBEI/II- line, and remained CA-type in WX9915-SBEI/II- line. These results provided some information for quality breeding and utilizations of rice with inhibition of SBE.


Assuntos
Enzima Ramificadora de 1,4-alfa-Glucana/antagonistas & inibidores , Endosperma/enzimologia , Oryza/enzimologia , Amido/química , Amilopectina/química , Amilopectina/metabolismo , Amilose/análise , Endosperma/efeitos dos fármacos , Oryza/química , Oryza/efeitos dos fármacos , Oryza/metabolismo , Amido/metabolismo
2.
Plant Physiol Biochem ; 142: 440-451, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31419646

RESUMO

Drought stress during the grain filling stage severely affects the quality and quantity of starch in rice grains. The enzymes such as ADP-glucose pyrophosphorylase (AGPase, EC 2.7.7.27) and starch synthase (SS, EC 2.4.1.21) are the key regulatory enzymes involved in the starch biosynthesis. In this study, the activity of the AGPase and starch synthase (SS) was correlated with the qualitative and quantitative parameters such as sucrose, starch, amylose, amylopectin, and resistant starch in leaves, roots, and grains of drought tolerant (N22) and drought susceptible (IR64) cultivars under applied water deficit stress (WDS). Drought stress enhanced the remobilization of stored starch from leaves to developing rice grains which was positively correlated with a decrease in the starch and starch synthase activity in leaves. Starch accumulation in developing grains was positively correlated with an increase in the AGPase and SS activity under drought. It was found that starch, amylopectin, and sucrose content in developing grains increased under water deficit stress (WDS), while amylose content decreased in both the varieties. However, in leaves, the SS activity decreased while AGPase activity was found to be increased under WDS in both varieties. Decreased starch content in matured grains was due to shortening of grain filling stage as drought stress caused early plant senescence. Yield reduction under drought was more in susceptible variety IR64 as compared to tolerant genotype N22.


Assuntos
Grão Comestível/metabolismo , Oryza/metabolismo , Amido/metabolismo , Amilopectina/metabolismo , Amilose/metabolismo , Desidratação , Grão Comestível/fisiologia , Oryza/fisiologia , Fotossíntese , Folhas de Planta/metabolismo , Sacarose/metabolismo
3.
Food Chem ; 295: 484-492, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31174785

RESUMO

The eating and cooking quality (ECQ) of stored rice grains is significantly affected by ageing, but the molecular mechanisms for this are not well understood. In the present study, changes of starch hierarchical structures and thermal properties of starch were investigated for rices with up to 12 months storage. For the first time, it was seen that storage resulted in molecular degradation of starch. This mainly involved shorter amylose chains and around (1 → 6)-α glycosidic branching points of amylopectin, which altered the crystalline structure. This resulted in lower gelatinization temperatures and enthalpy but higher crystalline heterogeneity. The ageing effect was varietally-dependent. The information obtained from this study offers improved molecular-level understanding of the effects of ageing process on rice cooking and eating qualities.


Assuntos
Armazenamento de Alimentos , Oryza/química , Amido/química , Amilopectina/química , Amilopectina/metabolismo , Amilose/química , Amilose/metabolismo , Cromatografia em Gel , Culinária , Gelatina/química , Gelatina/metabolismo , Microscopia Eletrônica de Varredura , Estrutura Molecular , Oryza/metabolismo , Espalhamento a Baixo Ângulo , Sementes/química , Amido/metabolismo , Fatores de Tempo , Difração de Raios X
4.
Food Chem ; 294: 248-259, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31126460

RESUMO

In this study, ultrasound was used to modulate the molecular structure of retrograded starch (RS3) responsible for the digestion properties, and the relationships among the ultrasonic power, molecular structure, and RS3 digestibility were revealed. Results revealed that the morphological characteristics of RS3 changed after ultrasonic treatment, and high power resulted in more compact block-shape structure. The results of particle size analysis, Fourier transform infrared spectroscopy, and X-ray diffraction showed that ultrasound decreased the long-range orders but increased the median particle size, short-range orders, and V-type polymorph of ultrasound-treated RS3. A decreased (double helix of amylopectin) and an increased (single helix of V-amylose) enthalpy change were observed by differential scanning calorimeter. Digestion profiles showed that some fractions of rapidly-digestible starch were converted into slowly-digestible starch and/or resistant starch. This study provides a potential approach to regulate the digestion of starch-rich foods with desired digestibility through ultrasonic treatment.


Assuntos
Amido/química , Amilopectina/química , Amilopectina/metabolismo , Amilose/química , Varredura Diferencial de Calorimetria , Conformação Molecular , Tamanho da Partícula , Solubilidade , Sonicação , Espectroscopia de Infravermelho com Transformada de Fourier , Amido/metabolismo , Difração de Raios X , alfa-Amilases/metabolismo
5.
Arch Anim Nutr ; 73(3): 194-207, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30864858

RESUMO

The present study was conducted to investigate the effects of dietary amylose and amylopectin ratio on growth performance, meat quality, postmortem glycolysis and muscle fibre type transformation of finishing pigs. Twenty-four barrows (Duroc × Landrace × Yorkshire) with an average initial body weight of 61.7 ± 2.01 kg were randomly assigned to four dietary treatments with amylose: amylopectin ratios of 1:1 (HD), 1:2 (MD), 1:3 (CD) and 1:4 (LD). The results showed that the average daily weight gain of finishing pigs tended to reduce with the ratio of amylose and amylopectin decreased (p = 0.09). Diet LD increased the pH24h value and decreased the shear force in longissimus dorsi (LM) compared with diet HD (p < 0.05). Diet LD decreased the lactate content and the HK-2 mRNA abundance and increased the mRNA abundance of ATP5B in LM compared with diet HD (p < 0.05). Higher mRNA abundance of MyHC I and lesser abundance of MyHC IIb in LM were found in pigs fed diet CD and LD than those fed diet HD (p < 0.05). Furthermore, pigs fed diet LD had higher mRNA abundances of PGC-1α and PPAR δ in LM than other groups (p < 0.05). These results suggested that diet with low amylose and amylopectin ratio could improve meat quality of finishing pigs via delaying muscle glycolysis capacity and shifting muscle fibre types.


Assuntos
Amilopectina/metabolismo , Amilose/metabolismo , Glicólise/fisiologia , Carne/análise , Fibras Musculares Esqueléticas/fisiologia , Sus scrofa/fisiologia , Amilopectina/administração & dosagem , Amilose/administração & dosagem , Ração Animal/análise , Fenômenos Fisiológicos da Nutrição Animal/efeitos dos fármacos , Animais , Dieta/veterinária , Relação Dose-Resposta a Droga , Glicólise/efeitos dos fármacos , Masculino , Fibras Musculares Esqueléticas/efeitos dos fármacos , Distribuição Aleatória , Sus scrofa/crescimento & desenvolvimento
6.
Food Chem ; 283: 375-380, 2019 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-30722886

RESUMO

In the present study, the effects of different irrigation levels (I50: 50%; I75: 75%; and I100: 100% of depleted water from field capacity) and nitrogen fertilizer concentrations (N1:100, N2:200 and N3:300 kg ha-1N) on starch properties (total, resistant and nonresistant starch, amylose-amylopectin ratios) and phytic acid content of maize were investigated. The single and interaction effects of the treatments and also influence of harvesting year were evaluated. Total starch accumulation changed with the treatments and the highest values were obtained from I50 and N3 combination and the lowest levels were recorded for I100-N1 and I100-N2 treatments. Amylopectin increased with the effect of irrigation, while the amylose decreased with the irrigation treatment in parallel to amylopectin content. Besides, the lowest phytic acid level was also obtained from the treatments of I100 and N1. This study would provide some valuable information for agricultural and nutritional applications of maize.


Assuntos
Fertilizantes/análise , Ácido Fítico/análise , Amido/química , Agricultura , Amilopectina/metabolismo , Amilose/metabolismo , Nitrogênio/química , Estações do Ano , Solo/química , Zea mays/química , Zea mays/metabolismo
7.
Food Chem ; 259: 89-98, 2018 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-29680067

RESUMO

Vibrational spectroscopic techniques were employed to predict the mechanism of starch hydrolysis based on structural changes during germination of paddy. The proposed mechanism for starch hydrolysis dealt with the synthesis of amylase at the onset of germination, depicting an increased intensity of spectral bands at amide I, II and III regions. The process commenced with the enzyme actions on skeletal mode of pyranose ring structure of glucose units followed by cleavage of the glycosidic linkage by the process of multiple and multi-chain attack resulting in decrease of the bands (400-900 cm-1). The increased intensity of the bands (1200-1500 cm-1) indicated the process of starch hydrolysis and formation of d-glucose. Multivariate calibration analysis (PCA and PLS) was employed to correlate Raman spectral data with biochemical changes during germination and to develop a calibration model. The model showed a high prediction ability with low root mean square error of prediction (RMSEP) (0.043-0.568).


Assuntos
Oryza/metabolismo , Análise Espectral Raman/métodos , Amido/química , Amilopectina/análise , Amilopectina/metabolismo , Amilose/análise , Amilose/metabolismo , Calibragem , Glucose/metabolismo , Hidrólise , Análise dos Mínimos Quadrados , Análise Multivariada , Análise de Componente Principal , Espectroscopia de Infravermelho com Transformada de Fourier , Amido/metabolismo , Vibração
8.
BMC Plant Biol ; 18(1): 59, 2018 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-29636002

RESUMO

BACKGROUND: Starch is the major component of cereal grains and is composed of essentially linear amylose and highly branched amylopectin. The properties and composition of starch determine the use and value of grains and their products. Starch synthase (SS) I, SSIIa, and SSIIIa play central roles in amylopectin biosynthesis. These three SS isozymes also affect seed development, as complete loss of both SSI and SSIIIa under reduced SSIIa activity in rice lead to sterility, whereas presence of minimal SSI or SSIIIa activity is sufficient for generating fertile seeds. SSs, branching enzymes, and/or debranching enzymes form protein complexes in cereal. However, the relationship between starch properties and the formation of protein complexes remain largely unknown. To better understand this phenomenon, properties of starch and protein complex formation were analyzed using developing mutant rice seeds (ss1 L /ss2a L /ss3a) in which all three major SS activities were reduced. RESULTS: The SS activity of ss1 L /ss2a L /ss3a was 25%-30% that of the wild-type. However, the grain weight of ss1 L /ss2a L /ss3a was 89% of the wild-type, 55% of which was starch, showing considerable starch synthesis. The reduction of soluble SS activity in ss1 L /ss2a L /ss3a resulted in increased levels of ADP-glucose pyrophosphorylase and granule-bound starch synthase I, which are responsible for substrate synthesis and amylose synthesis, respectively. Together, these features led to an increase in apparent amylose content (34%) in ss1 L /ss2a L /ss3a compared with wild-type (20%). Gel filtration chromatography of the soluble proteins in ss1 L /ss2a L /ss3a showed that the majority of the starch biosynthetic enzymes maintained the similar elution patterns as wild-type, except that the amounts of high-molecular-weight SSI (> 300 kDa) were reduced and SSIIa of approximately 200-300 kDa were present instead of those > 440 kDa, which predominate in wild-type. Immuno-precipitation analyses suggested that the interaction between the starch biosynthetic enzymes maybe reduced or weaker than in wild-type. CONCLUSIONS: Although major SS isozymes were simultaneously reduced in ss1 L /ss2a L /ss3a rice, active protein complexes were formed with a slightly altered pattern, suggesting that the assembly of protein complexes may be complemented among the SS isozymes. In addition, ss1 L /ss2a L /ss3a maintained the ability to synthesize starch and accumulated less amylopectin and more amylose in starch.


Assuntos
Oryza/enzimologia , Oryza/metabolismo , Sementes/enzimologia , Sementes/metabolismo , Sintase do Amido/metabolismo , Amido/metabolismo , Amilopectina/metabolismo , Amilose/metabolismo
9.
BMC Plant Biol ; 18(1): 9, 2018 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-29310584

RESUMO

BACKGROUND: Endosperm starch provides prime energy for cereal seedling growth. Cereal endosperm with repression of starch branching enzyme (SBE) has been widely studied for its high resistant starch content and health benefit. However, in barley and maize, the repression of SBE changes starch component and amylopectin structure which affects grain germination and seedling establishment. A high resistant starch rice line (TRS) has been developed through inhibiting SBEI/IIb, and its starch has very high resistance to in vitro hydrolysis and digestion. However, it is unclear whether the starch resists in situ degradation in seed and influences seedling growth after grain germination. RESULTS: In this study, TRS and its wild-type rice cultivar Te-qing (TQ) were used to investigate the seedling growth, starch property changes, and in situ starch degradation during seedling growth. The slow degradation of starch in TRS seed restrained the seedling growth. The starch components including amylose and amylopectin were simultaneously degraded in TQ seeds during seedling growth, but in TRS seeds, the amylose was degraded faster than amylopectin and the amylopectin long branch-chains with B-type crystallinity had high resistance to in situ degradation. TQ starch was gradually degraded from the proximal to distal region of embryo and from the outer to inner in endosperm. However, TRS endosperm contained polygonal, aggregate, elongated and hollow starch from inner to outer. The polygonal starch similar to TQ starch was completely degraded, and the other starches with long branch-chains of amylopectin and B-type crystallinity were degraded faster at the early stage of seedling growth but had high resistance to in situ degradation during TRS seedling growth. CONCLUSIONS: The B-type crystallinity and long branch-chains of amylopectin in TRS seed had high resistance to in situ degradation, which inhibited TRS seedling growth.


Assuntos
Amilopectina/metabolismo , Oryza/metabolismo , Plântula/metabolismo , Enzima Ramificadora de 1,4-alfa-Glucana/genética , Enzima Ramificadora de 1,4-alfa-Glucana/metabolismo , Oryza/genética , Oryza/crescimento & desenvolvimento , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plântula/crescimento & desenvolvimento
10.
Plant Physiol ; 176(1): 582-595, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29133372

RESUMO

Rice (Oryza sativa) endosperm is mainly occupied by homogeneous polygonal starch from inside to outside. However, morphologically different (heterogeneous) starches have been identified in some rice mutants. How these heterogeneous starches form remains unknown. A high-amylose rice line (TRS) generated through the antisense inhibition of starch branching synthase I (SBEI) and SBEIIb contains four heterogeneous starches: polygonal, aggregate, elongated, and hollow starch; these starches are regionally distributed in the endosperm from inside to outside. Here, we investigated the relationship between SBE dosage and the morphological architecture of heterogeneous starches in TRS endosperm from the view of the molecular structure of starch. The results indicated that their molecular structures underwent regular changes, including gradually increasing true amylose content but decreasing amylopectin content and gradually increasing the ratio of amylopectin long chain but decreasing the ratio of amylopectin short chain. Granule-bound starch synthase I (GBSSI) amounts in the four heterogeneous starches were not significantly different from each other, but SBEI, SBEIIa, and SBEIIb showed a gradually decreasing trend. Further immunostaining analysis revealed that the gradually decreasing SBEs acting on the formation of the four heterogeneous granules were mainly due to the spatial distribution of the three SBEs in the endosperm. It was suggested that the decreased amylopectin in starch might remove steric hindrance and provide extra space for abundant amylose accumulation when the GBSSI amount was not elevated. Furthermore, extra amylose coupled with altered amylopectin structure possibly led to morphological changes in heterogeneous granules.


Assuntos
Enzima Ramificadora de 1,4-alfa-Glucana/metabolismo , Grânulos Citoplasmáticos/enzimologia , Oryza/enzimologia , Plantas Geneticamente Modificadas/metabolismo , Amido/metabolismo , Amilopectina/química , Amilopectina/metabolismo , Amilose/metabolismo , Regulação para Baixo , Endosperma/enzimologia , Pleiotropia Genética , Isoenzimas/metabolismo , Proteínas de Plantas/metabolismo
11.
Food Chem ; 244: 136-142, 2018 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-29120761

RESUMO

Effect of debranching for a high-amylose starch (∼70% amylose) on its V-complex formation with stearic acid was examined. Gel-permeation chromatograms showed that amylopectin was degraded to smaller molecules as the debranching time increased from 6 to 24 h. Increased formation of debranched starch/stearic acid complexes (recovery yield of stearic acid from 45.17 to 89.31% and starch from 39.92 to 55.43%) was observed with increased debranching time (from 6 to 24 h) and complexation time (from 6 to 24 h). The X-ray diffraction patterns of the debranched starch/stearic acid complexes displayed a mixture of B-type and V-type patterns, with 2θ peaks at 7.6°, 13.1°, 17.2°, 20°, 21.6°, and 23.4°. The melting temperature and enthalpy changes of the debranched starch/stearic acid complexes were gradually enhanced with increasing debranching time. These results suggest that starch can be modified by debranching to produce a significant amount of debranched starch/stearic acid complexes.


Assuntos
Amido/metabolismo , Ácidos Esteáricos/metabolismo , Amilopectina/metabolismo , Amilose/análise , Amilose/metabolismo , Glicosídeo Hidrolases/metabolismo , Amido/química , Temperatura Ambiente , Termodinâmica , Difração de Raios X , Zea mays/química
12.
Food Funct ; 8(10): 3654-3663, 2017 Oct 18.
Artigo em Inglês | MEDLINE | ID: mdl-28914314

RESUMO

Consumers and the food industry are demanding healthier products. Expanded snacks with a high nutritional value were developed from different rice, pea and carob flour blends. The proximate composition, starch (total and resistant), amylose and amylopectin, dietary fiber (soluble and insoluble) contents, and the in vitro protein digestibility of different rice-legume formulations, were evaluated before and after the extrusion process. Compared with the corresponding non-extruded blends (control), the extrusion treatment did not change the total protein content, however, it reduced the soluble protein (61-86%), the fat (69-92%) and the resistant starch contents (100%). The total starch content of all studied blends increased (2-19%) after extrusion. The processing increased the in vitro protein digestibility, reaching values around 88-95% after extrusion. Total dietary fiber was reduced around 30%, and the insoluble fraction was affected to a larger extent than the soluble fraction by the extrusion process. Because of its balanced nutritional composition, high dietary fiber content, as well as low energy density, these novel gluten-free snack-like foods could be considered as functional foods and a healthier alternative to commercially available gluten-containing or gluten-free and low nutritional value snacks.


Assuntos
Fibras na Dieta/análise , Farinha/análise , Oryza/química , Ervilhas/química , Amilopectina/análise , Amilopectina/metabolismo , Culinária , Fibras na Dieta/metabolismo , Digestão , Glutens/análise , Glutens/metabolismo , Humanos , Valor Nutritivo , Oryza/metabolismo , Ervilhas/metabolismo , Lanches , Amido/análise , Amido/metabolismo
13.
BMC Plant Biol ; 17(1): 132, 2017 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-28764648

RESUMO

BACKGROUND: Although starch consists of large macromolecules composed of glucose units linked by α-1,4-glycosidic linkages with α-1,6-glycosidic branchpoints, variation in starch structural and functional properties is found both within and between species. Interest in starch genetics is based on the importance of starch in food and industrial processes, with the potential of genetics to provide novel starches. The starch metabolic pathway is complex but has been characterized in diverse plant species, including pea. RESULTS: To understand how allelic variation in the pea starch metabolic pathway affects starch structure and percent amylose, partial sequences of 25 candidate genes were characterized for polymorphisms using a panel of 92 diverse pea lines. Variation in the percent amylose composition of extracted seed starch and (amylopectin) chain length distribution, one measure of starch structure, were characterized for these lines. Association mapping was undertaken to identify polymorphisms associated with the variation in starch chain length distribution and percent amylose, using a mixed linear model that incorporated population structure and kinship. Associations were found for polymorphisms in seven candidate genes plus Mendel's r locus (which conditions the round versus wrinkled seed phenotype). The genes with associated polymorphisms are involved in the substrate supply, chain elongation and branching stages of the pea carbohydrate and starch metabolic pathways. CONCLUSIONS: The association of polymorphisms in carbohydrate and starch metabolic genes with variation in amylopectin chain length distribution and percent amylose may help to guide manipulation of pea seed starch structural and functional properties through plant breeding.


Assuntos
Amilose/metabolismo , Metabolismo dos Carboidratos/genética , Genes de Plantas , Ervilhas/metabolismo , Amido/metabolismo , Alelos , Amilopectina/metabolismo , Configuração de Carboidratos , Ervilhas/genética , Polimorfismo Genético , Amido/química
14.
MBio ; 8(4)2017 08 29.
Artigo em Inglês | MEDLINE | ID: mdl-28851850

RESUMO

In immunocompromised hosts, latent infection with Toxoplasma gondii can reactivate from tissue cysts, leading to encephalitis. A characteristic of T. gondii bradyzoites in tissue cysts is the presence of amylopectin granules. The regulatory mechanisms and role of amylopectin accumulation in this organism are not fully understood. The T. gondii genome encodes a putative glycogen phosphorylase (TgGP), and mutants were constructed to manipulate the activity of TgGP and to evaluate the function of TgGP in amylopectin storage. Both a stop codon mutant (Pru/TgGPS25stop [expressing a Ser-to-stop codon change at position 25 in TgGP]) and a phosphorylation null mutant (Pru/TgGPS25A [expressing a Ser-to-Ala change at position 25 in TgGp]) mutated at Ser25 displayed amylopectin accumulation, while the phosphorylation-mimetic mutant (Pru/TgGPS25E [expressing a Ser-to-Glu change at position 25 in TgGp]) had minimal amylopectin accumulation under both tachyzoite and bradyzoite growth conditions. The expression of active TgGPS25S or TgGPS25E restored amylopectin catabolism in Pru/TgGPS25A To understand the relation between GP and calcium-dependent protein kinase 2 (CDPK2), which was recently reported to regulate amylopectin consumption, we knocked out CDPK2 in these mutants. PruΔcdpk2/TgGPS25E had minimal amylopectin accumulation, whereas the Δcdpk2 phenotype in the other GP mutants and parental lines displayed amylopectin accumulation. Both the inactive S25A and hyperactive S25E mutant produced brain cysts in infected mice, but the numbers of cysts produced were significantly less than the number produced by the S25S wild-type GP parasite. Complementation that restored amylopectin regulation restored brain cyst production to the control levels seen in infected mice. These data suggest that T. gondii requires tight regulation of amylopectin expression for efficient production of cysts and persistent infections and that GP phosphorylation is a regulatory mechanism involved in amylopectin storage and utilization.IMPORTANCEToxoplasma gondii is an obligate intracellular parasite that causes disease in immune-suppressed individuals, as well as a fetopathy in pregnant women who acquire infection for the first time during pregnancy. This parasite can differentiate between tachyzoites (seen in acute infection) and bradyzoites (seen in latent infection), and this differentiation is associated with disease relapse. A characteristic of bradyzoites is that they contain cytoplasmic amylopectin granules. The regulatory mechanisms and the roles of amylopectin granules during latent infection remain to be elucidated. We have identified a role of T. gondii glycogen phosphorylase (TgGP) in the regulation of starch digestion and a role of posttranslational modification of TgGP, i.e., phosphorylation of Ser25, in the regulation of amylopectin digestion. By manipulating TgGP activity in the parasite with genome editing, we found that the digestion and storage of amylopectin due to TgGP activity are both important for latency in the brain.


Assuntos
Amilopectina/metabolismo , Encéfalo/parasitologia , Glicogênio Fosforilase/metabolismo , Toxoplasma/fisiologia , Amilopectina/genética , Animais , Diferenciação Celular , Glicogênio/metabolismo , Interações Hospedeiro-Patógeno , Camundongos , Fosforilação , Processamento de Proteína Pós-Traducional , Toxoplasma/enzimologia , Toxoplasmose Animal/parasitologia
15.
Plant Sci ; 262: 1-8, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28716405

RESUMO

Starch accounts for the majority of edible carbohydrate resources generated through photosynthesis. Amylopectin is the major component of starch and is one of highest-molecular-weight biopolymers. Rapid and systematic synthesis of frequently branched hydro-insoluble amylopectin and efficient accumulation into amyloplasts of cereal endosperm is crucial. The functions of multiple starch biosynthetic enzymes, including elongation, branching, and debranching enzymes, must be temporally and spatially coordinated. Accordingly, direct evidence of protein-protein interactions of starch biosynthetic enzymes were first discovered in developing wheat endosperm in 2004, and they have since been shown in the developing seeds of other cereals. This review article describes structural characteristics of starches as well as similarities and differences in protein complex formation among different plant species and among mutant plants that are deficient in specific starch biosynthetic enzymes. In addition, evidence for protein complexes that are involved in the initiation stages of starch biosynthesis is summarized. Finally, we discuss the significance of protein complexes and describe new methods that may elucidate the mechanisms and roles of starch biosynthetic enzyme complexes.


Assuntos
Grão Comestível/enzimologia , Grão Comestível/metabolismo , Amilopectina/genética , Amilopectina/metabolismo , Grão Comestível/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Sementes/enzimologia , Sementes/genética , Sementes/metabolismo , Amido/metabolismo , Triticum/enzimologia , Triticum/genética , Triticum/metabolismo
16.
Biochem Soc Trans ; 45(4): 885-893, 2017 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-28673938

RESUMO

Starch is the most widespread and abundant storage carbohydrate in plants and the main source of carbohydrate in the human diet. Owing to its remarkable properties and commercial applications, starch is still of growing interest. Its unique granular structure made of intercalated layers of amylopectin and amylose has been unraveled thanks to recent progress in microscopic imaging, but the origin of such periodicity is still under debate. Both amylose and amylopectin are made of linear chains of α-1,4-bound glucose residues, with branch points formed by α-1,6 linkages. The net difference in the distribution of chain lengths and the branching pattern of amylose (mainly linear), compared with amylopectin (racemose structure), leads to different physico-chemical properties. Amylose is an amorphous and soluble polysaccharide, whereas amylopectin is insoluble and exhibits a highly organized structure of densely packed double helices formed between neighboring linear chains. Contrarily to starch degradation that has been investigated since the early 20th century, starch production is still poorly understood. Most enzymes involved in starch growth (elongation, branching, debranching, and partial hydrolysis) are now identified. However, their specific action, their interplay (cooperative or competitive), and their kinetic properties are still largely unknown. After reviewing recent results on starch structure and starch growth and degradation enzymatic activity, we discuss recent results and current challenges for growing polysaccharides on granular surface. Finally, we highlight the importance of novel stochastic models to support the analysis of recent and complex experimental results, and to address how macroscopic properties emerge from enzymatic activity and structural rearrangements.


Assuntos
Modelos Moleculares , Plantas/metabolismo , Amido/metabolismo , Amilopectina/biossíntese , Amilopectina/química , Amilopectina/metabolismo , Amilose/biossíntese , Amilose/química , Amilose/metabolismo , Configuração de Carboidratos , Grânulos Citoplasmáticos , Glucanos/biossíntese , Glucanos/química , Glucanos/metabolismo , Hidrólise , Proteínas de Plantas/metabolismo , Plantas/enzimologia , Técnicas de Síntese em Fase Sólida/tendências , Solubilidade , Amido/biossíntese , Amido/química , Processos Estocásticos
17.
Int J Biol Macromol ; 105(Pt 1): 1001-1009, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28746886

RESUMO

The effects of the molecular structures of amylopectins on the enzymatic hydrolysis of waxy-rice amylopectin (WRA) and normal-rice amylopectin (NRA) were investigated. The results indicated that compared to NRA, WRA possessed larger chain length, number of chains, internal chain length, degree of polymerisation, interblock chain length, and lower degree of branching and short:long B-chains, which caused WRA was much less susceptible to enzymatic hydrolysis than NRA. The digestibility curves for WRA and NRA were well fitted by the first-order kinetic equation. WRA and NRA were hydrolyzed in two separates phases in the LOS plots. Whether using α-amylase alone, or together with amyloglucosidase, WRA and NRA exhibited different digestibility rates due to different chain structure of amylopectin. The low C1∞ values predicted that WRA and NRA would have little impact on blood glucose concentrations in the early digestion stage. HPLC results showed that G1∼G5 from WRA using α-amylase were lower than that from NRA, whereas from WRA were higher than that from NRA. There was the synergism between α-amylase and amyloglucosidase in glucose released from WRA and NRA.


Assuntos
Amilopectina/química , Glucana 1,4-alfa-Glucosidase/metabolismo , Oryza/química , alfa-Amilases/metabolismo , Amilopectina/metabolismo , Animais , Digestão , Hidrólise , Cinética , Coelhos
18.
Plant Mol Biol ; 94(4-5): 399-417, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28466433

RESUMO

The lengths of amylopectin-branched chains are precise and influence the physicochemical properties of starch, which determine starch functionality. Three major isozymes of starch synthases (SSs), SSI, SSII(a), and SSIII(a), are primarily responsible for amylopectin chain elongation in the storage tissues of plants. To date, the majority of reported rice mutants were generated using japonica cultivars, which have almost inactive SSIIa. Although three SSs share some overlapping chain length preferences, whether they complement each other remains unknown due to the absence of suitable genetic combinations of materials. In this study, rice ss1/SS2a/SS3a and SS1/SS2a/ss3a were newly generated, and the chain length distribution patterns of all the possible combinations of presence and absence of SSI, SSIIa, and SSIIIa activities were compared. This study demonstrated that SSIIa can complement most SSI functions that use glucan chains with DP 6-7 to generate DP 8-12 chains but cannot fully compensate for the elongation of DP 16-19 chains. This suggests that SSIIa preferentially elongates outer but not inner chains of amylopectin. In addition, the results revealed that neither SSI nor SSIIIa compensate for SSIIa. Neither SSI nor SSIIa compensate for elongation of DP >30 by SSIIIa. SSIIa could not resolve the pleiotropic increase of SSI caused by the absence of SSIIIa; instead, SSIIa further elongated those branches elongated by SSI. These results revealed compensatory differences among three major SS isozymes responsible for lengths of amylopectin branches.


Assuntos
Amilopectina/metabolismo , Regulação Enzimológica da Expressão Gênica/fisiologia , Regulação da Expressão Gênica de Plantas/fisiologia , Oryza/metabolismo , Proteínas de Plantas/metabolismo , Sintase do Amido/metabolismo , Amilopectina/química , Endosperma/química , Endosperma/metabolismo , Genótipo , Estrutura Molecular , Mutagênese Insercional , Oryza/genética , Amido/biossíntese , Sintase do Amido/classificação , Sintase do Amido/genética
19.
Appl Environ Microbiol ; 83(12)2017 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-28411221

RESUMO

Of the few predicted extracellular glycan-active enzymes, glycoside hydrolase family 13 subfamily 14 (GH13_14) pullulanases are the most common in human gut lactobacilli. These enzymes share a unique modular organization, not observed in other bacteria, featuring a catalytic module, two starch binding modules, a domain of unknown function, and a C-terminal surface layer association protein (SLAP) domain. Here, we explore the specificity of a representative of this group of pullulanases, Lactobacillus acidophilus Pul13_14 (LaPul13_14), and its role in branched α-glucan metabolism in the well-characterized Lactobacillus acidophilus NCFM, which is widely used as a probiotic. Growth experiments with L. acidophilus NCFM on starch-derived branched substrates revealed a preference for α-glucans with short branches of about two to three glucosyl moieties over amylopectin with longer branches. Cell-attached debranching activity was measurable in the presence of α-glucans but was repressed by glucose. The debranching activity is conferred exclusively by LaPul13_14 and is abolished in a mutant strain lacking a functional LaPul13_14 gene. Hydrolysis kinetics of recombinant LaPul13_14 confirmed the preference for short-branched α-glucan oligomers consistent with the growth data. Curiously, this enzyme displayed the highest catalytic efficiency and the lowest Km reported for a pullulanase. Inhibition kinetics revealed mixed inhibition by ß-cyclodextrin, suggesting the presence of additional glucan binding sites besides the active site of the enzyme, which may contribute to the unprecedented substrate affinity. The enzyme also displays high thermostability and higher activity in the acidic pH range, reflecting adaptation to the physiologically challenging conditions in the human gut.IMPORTANCE Starch is one of the most abundant glycans in the human diet. Branched α-1,6-glucans in dietary starch and glycogen are nondegradable by human enzymes and constitute a metabolic resource for the gut microbiota. The role of health-beneficial lactobacilli prevalent in the human small intestine in starch metabolism remains unexplored in contrast to colonic bacterial residents. This study highlights the pivotal role of debranching enzymes in the breakdown of starchy branched α-glucan oligomers (α-limit dextrins) by human gut lactobacilli exemplified by Lactobacillus acidophilus NCFM, which is one of the best-characterized strains used as probiotics. Our data bring novel insight into the metabolic preference of L. acidophilus for α-glucans with short α-1,6-branches. The unprecedented affinity of the debranching enzyme that confers growth on these substrates reflects its adaptation to the nutrient-competitive gut ecological niche and constitutes a potential advantage in cross-feeding from human and bacterial dietary starch metabolism.


Assuntos
Proteínas de Bactérias/metabolismo , Glucanos/metabolismo , Glicosídeo Hidrolases/metabolismo , Lactobacillus acidophilus/enzimologia , Amilopectina/química , Amilopectina/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Estabilidade Enzimática , Trato Gastrointestinal/microbiologia , Glucanos/química , Glicosídeo Hidrolases/química , Glicosídeo Hidrolases/genética , Humanos , Hidrólise , Cinética , Lactobacillus acidophilus/genética , Lactobacillus acidophilus/metabolismo , Especificidade por Substrato
20.
Int J Biol Macromol ; 101: 973-982, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28366860

RESUMO

Debranching of (1, 6)-α-linkages in starch is of great significance as it is widely used in different industries. In recent years, identifying a single potential enzyme that could debranch (1, 6)-α-linkages and cleave (1, 4)-α-linkages brought great interest as amylopectin possessed both (1, 4)-α- and (1, 6)-α-linkages. In the present study, a 64\,kDa exo-α-l, 4-glucosidase from Bacillus pseudofirmus 703, annotated as Amy112, was cloned and characterized. Biochemical analysis indicated that heterologous Amy112 expressed in Escherichia coli exhibited a high activity against amylopectin, with the optimal temperature and pH of 40°C and pH 7.0, respectively. Addition of K+ ions improved the amy112 activity by 12%, but Li+, Ca2+ and Mg2+ ions showed no significant effect. Amy112 sequence homology revealed that it belonged to glycoside hydrolase family 13, showing 65% identity with α-glucosidase GSJ from Geobacillus sp. HTA-462. This is the first report indicating that Amy112 from B. pseudofirmus 703 belongs to GH13 enzyme family, having both exo-α-1, 4-glucosidase and oligo-l, 6-glucosidase activities. However, no transglycosylation activity was detected in LC-MS analysis. Amy112 would be of great significance of being utilized to debranch starch in different industries in a cost effective manner.


Assuntos
Amilopectina/metabolismo , Bacillus/enzimologia , alfa-Glucosidases/química , alfa-Glucosidases/metabolismo , Sequência de Aminoácidos , Concentração de Íons de Hidrogênio , Cinética , Metais/farmacologia , Modelos Moleculares , Conformação Proteica , Solventes/farmacologia , Especificidade por Substrato , Temperatura Ambiente
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