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Studies on the chemical modification and potato (Solanum tuberosum) lectin and its effect on haemagglutinating activity.

ABSTRACT: 1. Modification of potato (Solanum tuberosum) lectin with acetic anhydride blocked 5.1 amino and 2.7 tyrosyl groups per molecule of lectin and decreased the haemagglutinating activity of the lectin. De-O-acetylation regenerated 2.0 of the tyrosyl groups and resulted in a recovery of activity. 2. Modification with citraconic anhydride or cyclohexane-1,2-dione did not greatly affect activity, although modification of amino and arginyl groups could be demonstrated. 3. Treatment with tetranitromethane nitrated 3.7 tyrosine residues per molecule of lectin with concomitant loss of activity. The presence of 0.1m-NN'N''-triacetylchitotriose (a potent inhibitor of the lectin) in the reaction medium protected all the tyrosyl residues from nitration and the lectin was fully active. 4. Modification of tryptophyl groups with 2-hydroxy-5-nitrobenzyl bromide and 2,3-dioxoindoline-5-sulphonic acid modified 0.9 and 2.6 residues per molecule of lectin respectively with a loss of activity in each case. Reaction of potato lectin with 2,3-dioxoindoline-5-sulphonic acid in the presence of inhibitor protected 2.4 residues of tryptophan from the reagent. Loss of haemagglutination activity was prevented under these conditions. 5. Reaction of carboxy groups, activated with carbodi-imide, with alpha-aminobutyric acid methyl ester led to the incorporation of 5.3 residues of the ester per molecule of lectin. Presence of inhibitor in this case, although protecting activity, did not prevent modification of carboxy groups; in fact an increase in the number of modified residues was seen. This effect could be imitated by performing the reaction in 8m-urea. In both cases the number of carboxy groups modified was close to the total number of free carboxy groups as determined by the method of Hoare & Koshland [(1967) J. Biol. Chem.242, 2447-2453]. Guanidination of lysine residues after carboxy-group modification gave less homoarginine than did the unmodified lectin under the same conditions, suggesting the formation of intramolecular cross-links during carbodi-imide activation. 6. It is suggested from the results presented that amino, arginyl, methionyl, histidyl and carboxyl groups are not involved in the activity of the lectin and that tyrosyl and tryptophyl groups are very closely involved. These findings are similar to those reported for other proteins that bind N-acetylglucosamine oligomers and also fit the general trend in other lectins.

SUBMITTER: Ashford D

PROVIDER: S-EPMC1163383 | biostudies-other | 1981 Nov

REPOSITORIES: biostudies-other

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Project description:BACKGROUND: Invertases are ubiquitous enzymes that irreversibly cleave sucrose into fructose and glucose. Plant invertases play important roles in carbohydrate metabolism, plant development, and biotic and abiotic stress responses. In potato (Solanum tuberosum), invertases are involved in 'cold-induced sweetening' of tubers, an adaptive response to cold stress, which negatively affects the quality of potato chips and French fries. Linkage and association studies have identified quantitative trait loci (QTL) for tuber sugar content and chip quality that colocalize with three independent potato invertase loci, which together encode five invertase genes. The role of natural allelic variation of these genes in controlling the variation of tuber sugar content in different genotypes is unknown. RESULTS: For functional studies on natural variants of five potato invertase genes we cloned and sequenced 193 full-length cDNAs from six heterozygous individuals (three tetraploid and three diploid). Eleven, thirteen, ten, twelve and nine different cDNA alleles were obtained for the genes Pain-1, InvGE, InvGF, InvCD141 and InvCD111, respectively. Allelic cDNA sequences differed from each other by 4 to 9%, and most were genotype specific. Additional variation was identified by single nucleotide polymorphism (SNP) analysis in an association-mapping population of 219 tetraploid individuals. Haplotype modeling revealed two to three major haplotypes besides a larger number of minor frequency haplotypes. cDNA alleles associated with chip quality, tuber starch content and starch yield were identified. CONCLUSIONS: Very high natural allelic variation was uncovered in a set of five potato invertase genes. This variability is a consequence of the cultivated potato's reproductive biology. Some of the structural variation found might underlie functional variation that influences important agronomic traits such as tuber sugar content. The associations found between specific invertase alleles and chip quality, tuber starch content and starch yield will facilitate the selection of superior potato genotypes in breeding programs.

Project description:BACKGROUND:The natural variation of starch phosphate content in potatoes has been previously reported. It is known that, in contrast to raw starch, commercially phosphorylated starch is more stable at high temperatures and shear rates and has higher water capacity. The genetic improvement of phosphate content in potato starch by selection or engineering would allow the production of phosphorylated starch in a natural, environmentally friendly way without chemicals. The aim of the current research is to identify genomic SNPs associated with starch phosphorylation by carrying out a genome-wide association study in potatoes. RESULTS:A total of 90 S. tuberosum L. varieties were used for phenotyping and genotyping. The phosphorus content of starch in 90 potato cultivars was measured and then statistically analysed. Principal component analysis (PCA) revealed that the third and eighth principal components appeared to be sensitive to variation in phosphorus content (p = 0.0005 and p = 0.002, respectively). PC3 showed the correlation of starch phosphorus content with allelic variations responsible for higher phosphorylation levels, found in four varieties. Similarly, PC8 indicated that hybrid 785/8-5 carried an allele associated with high phosphorus content, while the Impala and Red Scarlet varieties carried alleles for low phosphorus content. Genotyping was carried out using an Illumina 22 K SNP potato array. A total of 15,214 scorable SNPs (71.7% success rate) was revealed. GWAS mapping plots were obtained using TASSEL based on several statistical models, including general linear models (GLMs), with and without accounting for population structure, as well as MLM. A total of 17 significant SNPs was identified for phosphorus content in potato starch, 14 of which are assigned to 8 genomic regions on chromosomes 1, 4, 5, 7, 8, 10, and 11. Most of the SNPs identified belong to protein coding regions; however, their allelic variation was not associated with changes in protein structure or function. CONCLUSIONS:A total of 8 novel genomic regions possibly associated with starch phosphorylation on potato chromosomes 1, 4, 5, 7, 8, 10, and 11 was revealed. Further validation of the SNPs identified and the analysis of the surrounding genomic regions for candidate genes will allow better understanding of starch phosphorylation biochemistry. The most indicative SNPs may be useful for developing diagnostic markers to accelerate the breeding of potatoes with predetermined levels of starch phosphorylation.

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Studies on the chemical modification and potato (Solanum tuberosum) lectin and its effect on haemagglutinating activity.

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