pericarp tissue from red ripe fruit of the processing tomato , heinz 2605 , was soaked for 3 hours in 0 . 1 % sodium metabisulfite at 2 ° c . and than rinsed for 48 hours in cold , running water . excess water was removed from the residue before it was homogenized in 100 mm sodium citrate , 5 . 5 m nacl , 40 mm β - mercaptoethanol , ph4 . 0 ( equal w / v ). the homogenate was left overnight at 220 c . before centrifuging at 16 , 300 g for 50 min . the supernatant was concentrated to 200 ml by ultrafiltration ( sty30 cartridge , amicon scientific ) diluted with 100 mm citrate ph4 . 0 to reduce the nacl concentration to 200 mm and reconcentrated . the concentrate was clarified by centrifuging at 27 , 000 g for 60 min , and applied to a s - sepharose ff column ( pharmacia lkb biotechnology ) 25 × 250 mm , equilibrated with 100 mm citrate , 200 mm nacl , 0 . 1 mm dtt , ph4 . 0 . bound proteins were eluted with a concave gradient to 1500 mm nacl in 100 mm citrate , 0 . 1 mm dtt , ph4 . 0 . pg2 was the first peak of pg activity to elute and pg1 was the second peak . active fractions of pg1 were pooled , concentrated by ultrafiltration , dialysed against 50 mm phosphate , 200 mm nacl , 0 . 1 mm dtt , ph6 . 0 ( start buffer ) and applied to a fplc mono s hr 10 / 10 column ( pharmacia ) equilibrated with the start buffer . the enzyme was eluted using a linear gradient of nacl to 1000 mm and loaded on to a concanavalin a - sepharose 4b ( pharmacia ) column , 25 × 100 mm in 50 mm sodium phosphate buffer , 500 mm nacl , 1 mm mgcl 2 , 1 mm cacl 2 ph6 . 0 . unbound proteins were washed from the column before pg1 was eluted using a gradient of buffer containing from 0 - 200 mm α - methylamannoside . active fractions were pooled and concentrated ; 2 ml was applied to a column of sephacryl 300 hr ( pharmacia ), 25 × 800 mm , in 50 mm phosphate , 500 mm nacl , 0 . 1 mm dtt ph6 . 0 . fractions with pg1 activity were concentrated by ultrafiltration , diluted to 200 mm nacl and fractionated on a mono s hr 5 / 5 column using a linear gradient of nacl in 50 mm phosphate , 0 . 1 mm dtt ph6 . 0 . pg1 was incubated in dissociating buffer ( 50 mm phosphate , 75 mm nacl , 10 mm dtt , 6 m urea , ph6 . 0 ) overnight in an atmosphere of nitrogen at 20 ° c . subunits were separated on a mono s hr 5 / 5 column using a linear gradient of nacl to 500 mm in 50 mm phosphate , 6 m urea , 1 mm dtt ph6 . 0 . the separate subunits were concentrated by ultrafiltration using a ym10 ( amicon scientific ) membrane and rechromatographed under the same conditions , except at ph4 . 0 . prior to amino acid analysis or protein sequencing , proteins were reduced and carboxymethylated as described by sheehy et al . ( 1987 ). proteins were hydrolysed in 6 m hcl for 24 h in a nitrogen atmosphere . the phenylisothiocyanate derivatives of the released amino acids were analysed by the method of heinrikson and meredith ( 1984 ). n - terminal protein sequences were determined on a gas phase protein sequencer ( applied biosystems model 470a ). n - terminal sequencing on pg1 protein was carried out after carboxymerthylation by the methods described by sheehy et al ( 1987 ). the sequence of the second subunit was deduced from the double sequence of the pg1 protein and the known n - terminal sequences of the pg2 subunit . the n - terminal sequences were confirmed by sequencing the subunits individually after they had been separated by ion - exchange chromatography in 6 m urea . the second or β - subunit was carboxymethylated and , suspended in 0 . 1 m tris , 0 . 1 % sds ph8 . 35 digested with endoproteinase lys - c ( boehringer -- mannheim gmb h3 ) at 40 ° c . for 2 h . after centrifuging , peptides in the supernatant were separated using a brownlea c18 rp , 5u , 4 . 6 - 250 mm column and a gradient of 0 - 2 h 70 % acetonitrile in 0 - 1 % tfa . pg2 and the beta - subunit can combine in vitro to form pg1 or a pg1 - like compound ( pressey , 1988 ; knegt et al ., 1988 ). this raises the possibility that pg1 in extracts results from the co - extraction of the two polypeptides and their combination in the extraction fluid ( pressey , 1988 ). critical examination of the evidence for and against the existence of pg1 in situ in cell walls leads to the conclusion that the evidence is ambiguous . this is the case when the tissue is disrupted , or extracts are made . since pg1 is more resistant to heat than pg2 , this property can be used to investigate the presence of pg1 in tissues provided the heat can be applied without the tissue losing its structural integrity . the inactivation of the polygalacturonase enzyme activity was investigated in intact fruit heated to 60 ° c . a biphasic decay in activity was measured exactly as predicted from the proportions of pg1 and pg2 in extracts of the tissues ( fig5 ). this is strong evidence that pg1 exists as such in the tissue and that polygalacturonase activity in situ is mediated by an enzyme with the characteristics of pg1 . antibodies were raised in new zealand white rabbits to purified pg1 protein . each rabbit was injected subcutaneously with 1 mg of antigen emulsified with freund &# 39 ; s complete adjuvant . booster injections were made after 28 days , and blood samples were taken after a further 7 days . the blood was clotted and centrifuged , and the serum treated with ammonium sulphate to 40 % of saturation to remove albumins . precipitation was repeated once and the supernatant dialysed against phosphate buffered saline . the serum was affinity purified ( goding , 1986 ) by binding to pg2 coupled to sepharose 4b ( pharmacia ). unbound igg was washed from the column with phosphate - buffered saline and the bound fraction with 0 . 1 m acetic acid ; the bound fractionj was neutralised and dialysed against phosphate - buffered saline . the process of binding to a pg2 affinity column and elution was repeated once . the affinity - purified anti - pg1 serum recognised native pg1 but not pg2 protein ; on western blots ( kyhse - anderson , 1984 ; bjerrum and schafer - nielsen , 1986 ) it recognised the beta subunit . total protein extracts were made from tomato tissues in 50 mm sodium phosphate , 5 % sds , 6 m urea , 5 % glycarol , 10 % beta - mercaptoethanol ph 6 . 0 and the extracted proteins separated through polyacrylamide gels in the presence of sds . the separated proteins were blotted to nitrocellulose and probed with the purified anti - pg1 antisera . the beta - subunit antigen was detected in fruit pericarp tissue , but not in locule tissue or non - fruit tissues . in fruit pericarp , the antigen was detected 30 days but not 15 days post - anthesis . fig6 shows its concentration relative to dry matter in the pericarp of developing fruit of the cultivar de ruiter 83g38 . by avoiding ( nh 4 ) 2 so 4 precipitation and dialysis in buffers of low ionic strength , pg1 was purified from fruit of the processing line heinz 2605 ( table 1 ). overall purification was about 109 - fold and the yield about 10 %. the strong affinity of pg1 for cation exchange columns and for the lectin , concanavalin a , aided purification . purified pg1 , when reduced with β - mercaptoethanol and denatured with sds separated into three polypeptides of mr 45 k , 43 k and 39 k as estimated by sds - polyacrylamide electrophoresis ( fig1 ). a similar observation was made by moshrefi and luh ( 1983 ). the 45 k and 43 k , but not the 38 k polypeptides bound antiserum to pg2a , indicating that the smaller polypeptides was a distinct and different subunit of the pg1 molecule ( fig1 ). the smaller 39 k subunit , here termed the β - subunit , was shown to be chemically distinct . purified samples of pg1 and pg2 were dialyzed against nh 4 hco 3 , lyophilized and analyzed for carbohydrate ( table 2 ). both proteins contained glucosamine , mannose , xylose and fucose , but relative to the polypeptide , the carbohydrate content of pg1 was higher ( 18 . 9 %) than that of pg2 ( 11 . 2 %). since the larger subunits of pg1 are equivalent in size to pg2a and pg2b , the extracarbohydrate in pg1 is likely to be within the β - subunit . it was confirmed that the β - subunit stained intensely with schiff &# 39 ; s reagent ( result not shown ). the uv spectra of purified samples of pg1 and pg2 are shown in fig2 . pg1 has an absorption maximum about 275 nm and pg2 about 280 nm indicating that pg1 has the higher ratio of tyrosine to tryptophane . the relatively high content of tyrosine in pg1 was confirmed by amino acid analysis ( table 3 ). pg1 and pg2 are separable by chromatofocusing ( fig3 ) and it was confirmed ( ali and brady , 1982 ) that , although pg1 elutes after pg2 during cation exchange chromatography , pg1 has the lower isoelectric point . this suggested that the β - subunit is more acidic than pg2 . when pg1 was denatured in 6 m urea , 50 mm sodium phosphate , 75 mm nacl , 10 mm dtt ph6 . 0 , two polypeptides were separable by cation exchange chromatography ( fig4 ). the less tightly bound polypeptide corresponded to the β - subunit and the more tightly bound to pg2 as shown by sds - polyacrylamide gel electrophoresis ( fig4 ). the spectrum of the recovered presumptive β - subunit ( fig2 ) and its amino acid analysis ( table 3 ) showed a high content of tyrosine . on the basis of the amino acid analysis and the apparent molecular size , it was suggested that the β - subunit polypeptide was of mr 28 , 120 and contained 21 moles / mole of tyrosine . spectral analysis of the peptide in guanidine - hcl ( edelhoch , 1967 ) showed the presence of 1 mole / mole of tryptophane giving the polypeptide of mr of 28 , 342 . allowing that pg1 contains 1 mole each of pg2 and the β - subunit , it is calculated from the analysis in tables 2 and 3 , that the β - subunit has 27 . 9 % by weight of carbohydrate and a mr of 39 , 250 . pg2 had a polypeptide weight of 40 , 279 ( sheehy et al . 1987 ) and , from table 2 , 5075 g per mole of carbohydrate to give a subunit of 45 , 354 . again assuming 1 mole / mole of each subunit , the molecular mass of pg1 would be 84 , 600 ( table 3 ). the assumption of 1 mole each of the β - subunit and of pg2 in pg1 is not immediately consistent with the 3 subunits shown in fig1 . if the 43 k and 45 k components of pg1 represent the pg2a and pg2b forms of pg2 ( ali and brady 1982 ), and their recognition by specific antisera would suggest this , then it seems likely that pg1 is a population of related molecules containing either pg2a or pg2b linked with the β - subunit . the evidence is not entirely consistent with pg1 having a 1 : 1 mix of the β - subunit of pg2 . based on the tyrosine , tryptophane and cysteine contents , an absorption ratio , 280 nm to 288 nm of 1 . 74 was expected for pg1 but a ration of 1 . 695 was observed . recoveries of the subunits after cation exchange chromatography in urea was close to a molar ratio of 1 . 2 : 1 for pg2 : β - subunit . it is clear that pg2 and the β - subunit are major components of pg1 but more detailed analyses are required to define the structure . the n terminal of the pg2 subunit of pg1 was identical to that already described for the free pg2 protein ( sheehy et al , 1987 ). the n - terminal sequence of the second or β - subunit was determined by difference from the sequences from the composite pg1 , and from the unique sequence of the separated β - subunit . from several analysis the sequence emerged as : the latter sequence is of particular interest because of its homology with the 250 - 260 region of pg2 sequence - i - g - a - e - n - g - v - r - i - k - t - w - q there is much circumstantial evidence for the thesis that the pg1 isoform of endopolygalacturonase is involved in the initial catabolism of pectic polymers in ripening tomato fruits . the pg1 isoform is the only form of the enzyme recovered from tissue when softening is initiated . the present inventors have demonstrated that pg1 ( more than pg2 ) has the capacity , in vitro at least , to hydrolyse polygalacturonic substances in the low ionic conditions that may prevail in the fruit apoplast . ` nr ` mutant fruit and pg transgenic ` rin ` fruit have pectins with modified solubility ( seymour et al . 1987 ; giovannoni et al . 1989 ) and pg exclusively as the pg1 isomer ( tucker et al . 1980 ; bennett , private communication ; brady et al . 1983 ). homozygous nor and rin fruit have no pg and the pectin solubility remains unchanged ( seymour et al . 1987 ). circumstantially rhen , there is a link between the presence of pg1 and changes in the pectin polymers , and pg1 has the capacity to modify extracted pectin polymers . this circumstantial association is well short of proof that pg1 occurs and is active in vivo , but is sufficient to lead to an enquiry of the nature of the pg1 and the significance of the differences between pg1 and pg2 . since characterization of the pg1 converting factor ( s ) has proved elusive the present inventors have dissociated the native pg1 protein , isolated and partially characterised its subunits . characterising the actual components of pg1 will lead towards identifying the putative converting factor ( s ) and elucidation of the process of pg1 formation . the present inventors have successfully isolated and partially characterised the β - subunit of pg1 . they have also added to the evidence that the pg2 - like subunits of pg1 are indeed identical to pg2 by establishing that n - terminal sequence of pg2 is present in pg1 . the other major component of pg1 , the β - subunit , is a heavily - glycosylated , rather acidic polypeptide rich in glycine and tyrosine . the carbohydrate is present as n - linked n - acetylglucosamine , mannose , fucosa and xylose chains such as are common in plant glycoproteins ( ashfor , 1987 ). although it is necessary to have reducing agents present to separate the pg1 subunits in dissociating solvents , there is as yet no evidence that disulfide bonds couple the subunits . indeed the presence of cysteine residues in the β - subunit has not been demonstrated . sequence information on the β - subunit polypeptide is presented here for the first time . a molecular characterisation of the β - subunit should lead to probes that will enable its role through development to be evaluated and any role it has in siting or regulating pg action to be revealed . the sequence of the pg subunit opens the way to the isolation and sequencing of its gene . from the amino acid sequence a putative nucleic acid ( rna or dna ) can be synthesised and used ( a ) as a probe for use on cloned dna towards isolating the gene and ( b ) as template for use , with rna preparations , for production of cdna using the pcr technique . recovery of either gene or cdna segments would rapidly lead to an extension of the sequence information with the ability to alter gene expression by antisense suppression of the steady state mrna level in fruit . it may also be possible to use the putative nucleic acid sequence ( derived from the partial protein sequence ) directly as an antisense agent . for example , antisense genes may include sequence ( s ) coding for catalytic region ( s ) of riboendonucleases . the introduction of the antisense gene into tomato fruit is a well established technique ( sheehy et al . 1988 ; smith et al . 1988 ; hamilton et al 1990 ). the antisense gene is formulated behind a viral promoter . it is introduced into tomato cotyledon tissue via a transformation vector utilising agrobacterium tumefaciens ( fillatti et al 1987 ). plants are regenerated from the transformed tissue . the viral promoter ensures that the antisense element is expressed strongly and contitutively in the regenerated plant . alternatively , if expression is to be limited for example to fruit , a tissue specific promoter ( giovannoni et al 1989 ) may be used . the catalytic subunit of tomato fruit endopolygalacturonase has been suppressed by the antisense approach ( sheehy et al , 1988 , smith et al . 1988 ) with positive effects on fruit quality , especially for processing . when the total amount of endopolygalacturonase is limited , most of it is in the thermostable pg1 form . to prevent this enzyme causing an excessive degradation of pectin during processing , a &# 34 ; hot break &# 34 ; procedure must be used with attendant increases in energy costs . if the content of pg1 could be reduced , the &# 34 ; cold break &# 34 ;, low cost processing system would be applicable . use of the pg1 second subunit gene in the antisense form should result in a reduction in pg1 with a consequential improvement in the processed fruit . it is possible that the unprocessed fruit would retain firmness for a longer period for there is some evidence that pg1 is the form of endopolygalacturonase that is active within the fruit ( ostaryoung et al , 1990 ). it will be appreciated by persons skilled in the art that numerous variations and / or modifications may be made to the invention as shown in the specific embodiments without departing from the spirit or scope of the invention as broadly described . the present embodiments are , therefore , to be considered in all respects as illustrative and not restrictive . table 1__________________________________________________________________________purification of pg1 from ripe tomato fruit pg1 specific purification protein activity yield activityfraction ( mg ) ( ukat ) (%) ( ukat mg - 1 ) factor__________________________________________________________________________0 . 3m nacl extract 8800 0 0 0 1 . sup . a1 . 5m nacl extract 1278 109 100 0 . 085 8s - sepharose , ph4 141 68 62 . 268 24mono s , ph6 50 46 42 . 92 84con a 38 41 38 1 . 08 98sephacryl 300 hr 25 28 26 1 . 12 102mono s , ph6 10 12 11 1 . 2 109__________________________________________________________________________ periocarp weight fractionated 5 kg . sup . a from tissue , purification is 109 fold , because 8 . 8 g of protein without pg1 activiity is discarded in the preliminary extractions . table 2______________________________________ sample pg1 pg2component ( ug / mg polypeptide **) ______________________________________fucose 28 . 3 ± 0 . 9 8 . 1 ± 0 . 1xylose 27 . 7 ± 0 . 6 10 . 6 ± 0 . 4mannose 91 . 0 ± 1 . 7 44 . 0 ± 0 . 2x * 15 . 0 ± 0 . 2glucosamine 71 . 7 ± 3 . 1 62 . 4 ± 3 . 5______________________________________ * this unknown has a retention time , relative to inositol of 0 . 64 ** measured spectrophotometrically , allowing for differing extinction coefficients . figures are means with half the range from two independent analyses . table 3______________________________________amino acids in pg2a , pg1 and pg1 β - sub - unit pg2a . sup . 2molar ratios moles per mole of isoleucine ( moles per mole polypeptide ) β - sub - pg2a pg1 unit pg2a . sup . 2 β - subunit . sup . 3______________________________________asp ( n ) 1 . 28 . sup . 1 ( 1 . 50 ). sup . 4 2 . 54 . sup . 1 7 . 23 . sup . 1 54 52glu ( n ) 0 . 85 ( 0 . 98 ) 1 . 09 2 . 76 34 20ser 0 . 99 ( 1 . 08 ) 1 . 53 3 . 26 39 25gly 0 . 93 ( 0 . 86 ) 1 . 85 5 . 13 31 38his 0 . 16 ( 0 . 16 ) 0 . 29 0 . 73 6 6arg 0 . 22 ( 0 . 19 ) 0 . 33 0 . 86 7 6thr 0 . 53 ( 0 . 58 ) 1 . 18 2 . 79 21 21ala 0 . 64 ( 0 . 64 ) 0 . 96 2 . 49 23 19pro 0 . 49 ( 0 . 36 ) 0 . 66 1 . 26 13 10tyr 0 . 21 ( 0 . 28 ) 0 . 77 2 . 79 10 21val 0 . 81 ( 0 . 86 ) 1 . 07 1 . 63 31 13met 0 . 10 ( 0 . 08 ) 0 . 09 0 . 06 3 1cys 0 . 26 ( 0 . 39 ) 0 . 31 nd 14ile 1 . 00 ( 1 . 00 ) 1 . 00 1 . 00 36 7leu 0 . 51 ( 0 . 47 ) 0 . 57 0 . 26 17 2phe 0 . 45 ( 0 . 36 ) 0 . 42 0 . 25 13 2lys 0 . 92 ( 0 . 72 ) 1 . 07 1 . 77 26 14trp ( 0 . 11 ) 4 . sup . 11 . sup . 5______________________________________ . sup . 1 by direct analysis . sup . 2 from the sequence ( sheehy et al . 1987 ) . sup . 3 assumes polypeptide molecular mass of 27 , 342 for subunit . sup . 4 calculated from the sequence ( sheehy et al . 1987 ) . sup . 5 measured spectrophotometrically ali , z . m . and brady , c . j . purification and characterization of the polygalacturonases of tomato fruits . aust . j . plant physiol . 9 , 155 1982 . ashford , d ., dwek , r . a ., welply , j . k ., amatayakul , s ., homans , s . w ., lis , h ., taylor , g . n ., sharon , n . and rademacher , t . w . the β - 1 - 2d - xylose and α - 1 - 3 - 1 - 3 - l - fucose substituted n - linked oligosaccharides from erythrina cristacalli lectin . isolation , characterization and comparison with other legume lectins . eur . j . biochem . 166 , 311 , 1987 . bird , c . r ., smith , c . j . s ., ray , j . a ., moureau , p ., bevan , m . w ., bird , a . s ., hughes , s ., morris , p . c ., grierson , d . and schuch , w . the tomato polygalacturonase gene and ripening -- specific expression in transgenic plants . plant mol . biol . 11 , 651 , 1988 . bjerrum , o . j ., and schafer - nielsen , c . ( 1986 ) buffer systems and transfer parameters for semidry electroblotting with horizontal apparatus . in dunn , m . j . ( ed . ): electrophoresis &# 39 ; 86 . weinheim : vch , pp , 39 - 41 . brady , c . j ., macalpine , g ., mcglasson , w . b . and ueda , y . polygalacturonase in tomato fruits and the induction of ripening . aust . j . plant physiol . 9 , 171 , 1982 . brady , c . j ., meldrum , s . k ., mcglasson , w . b . and ali , z . m . differential accumulation of the molecular forms of polygalacturonase in tomato mutants . j . food biochem . 7 , 7 , 1983 . brady , c . j ., mcglasson , w . b . and speirs , j . ( 1987 ). the biochemistry of fruit ripening . in : tomato biotechnology . plant biology 4 , 279 , 1987 . ( eds d . j . nevins , r . a . jones ) ( alan r . liss inc : n . y .). dellapenna , d ., alexander , d . c . and bennett , a . b . molecular cloning of tomato fruit polygalacturonase : analysis of polygalacturonase mrna levels during ripening . proc . natl acad . sci . usa 83 , 6420 1986 . dellapenna , d ., lincoln , j . e ., fischer , r . l . and bennett , a . b . transcriptional analysis of polygalaturonase and other ripening associated genes in rutgers , rin nor and nr tomato fruit . plant physiol . 90 , 1372 , 1989 . edelhoch , h . spectroscopic determination of tryptophan and tryosine in proteins . biochemistry 6 , 1946 , 1967 . fillatti , j . j ., kiser , j ., rose , b . and commie , l ., efficient transformation of tomato and the introduction and expression of a gene for herbicide tolerance . in tomato biotechnology , d . j . nevis and r . a . jones , eds . ( n . y . : alan r . liss ) 199 , 1987 . giovannoni , j . j ., dellapenna , d ., bennett , a . b . and fisher , r . l . expression of a chimeric polygalacturonase gene in transgenic rin ( ripening inhibitor ) tomato fruit results in polyuronide degradation but not fruit softening . the plant cell 1 , 53 , 1989 . goding , j . w . ( 1986 ). monoclonal antibodies : principles and practice . sydney : academic press . grierson , d . gene expression in ripening tomato fruit . critical reviews in plant science . 3 , 113 , 1985 . grierson , d ., tucker , g . a . keen , j ., ray , j ., bird , c . r . and schuch , w . sequencing and identification of a cdna clone for tomato polygalacturonase . nucleic acids res . 14 , 8595 , 1986 . hamilton , a . j ., lycett , g . w . and grierson , d ., antisense gene that inhibits synthesis of the hormone ethylene in transgenic plants . nature 346 , 284 , 1990 . heinrickson , r . l . and meredith , s . c . amino acid analysis by reverse phase high performance liquid chromatography : precolumn derivatization with phenylisothiocyante . anal . biochem . 136 , 65 , 1984 . knegt , e ., varmeer , e . and bruinsma , j . conversion of the polygalacturonase isoenzymes from ripening tomato fruits . physiol . plant 72 , 108 , 1988 . khyse - anderson , j . ( 1984 ) electroblotting of multiple gels : a simple apparatus without buffer tank for rapid transfer of proteins from polyacrylamide to nitrocellulose . j . biochem . and biophys . methods 10 , pp 203 - 209 . maunders , m . j ., holdsworth , m . j ., slater , a . k ., knapp , j . e ., bird , c . r ., schuch , w . and grierson , d . ethylene stimulates the accumulation of ripening related messenger rnas in tomatoes . plant cell environ . 10 , 177 , 1987 . moshrefi , m . and luh , b . s . carbohydrate composition and electrophoretic properties of tomato polygalacturonase isoenzymes . eur . j . biochem . 135 , 511 , 1983 . osteryoung , k . w ., toenjes , k ., hall , b ., winkler , v . and bennett , a . a . analysis of tomato polygalacturonase expression in transgenic tobacco . the plant cell , 2 , ( 1239 ), 1990 . pressey , r . purification and characterization of tomato polygalacturonase converter . eur . j . biochem . 144 , 217 , 1984 . pressey , r . changes in polygalacturonase isoenzymes and converter in tomatoes during ripening . hort science 21 , 1183 , 1986 . pressey , r . reevaluation of the changes in polygalacturonases in tomatoes during ripening . plants 174 , 39 , 1988 . sawamura , m ., knegt , e . and bruinsma , j . levels of endogenous ethylene , carbon dioxide and soluble pectin and activities of pectin methylesterase and polygalacturonase in ripening of tomato fruits . plant cell physiol . 19 , 1061 , 1978 . seymour , g . b ., harding , s . e ., taylor , a . j ., hobson , g . e . and tucker , g . a . polyuronide solubilization during ripening of normal and mutant tomato fruit . phytochemistry 26 , 1871 , 1987 . sheehy , r . e ., pearson , j ., brady , c . j . and hiatt , w . r . molecular characterization of tomato fruit polygalacturonase . molecular and general genetics 208 , 30 , 1987 . sheehy , r ., kramer , m . and hiatt , w . r . reduction of polygalacturonase activity in tomato fruit by antisense rna . proc . natl . acad . sci . usa 85 , 8805 , 1988 . smith , c . j ., watson , c . f ., ray , j ., bird , r . c ., morris , p . c ., schuch , w . and grierson , d . antisense rna inhibition of polygalacturonase gene expression in transgenic tomatoes . nature 334 , 724 , 1988 . tucker , g . a ., robertson , n . g . and grierson , d . changes in polygalacturonase isoenzymes during the ripening of normal and mutant tomato fruit . eur . j . biochem . 112 , 119 , 1980 . tucker , g . a ., robertson , n . g . and grierson , d . the conversion of tomato - fruit polygalacturonase isoenzyme 2 into isoenzyme 1 in vitro . eur . j . biochem . 115 , 87 , 1981 . tucker , g . a . and grierson , d . synthesis of polygalacturonase during tomato fruit ripening . planta 155 , 64 , 1982 . __________________________________________________________________________sequence listing ( 1 ) general information :( iii ) number of sequences : 6 ( 2 ) information for seq id no : 1 :( i ) sequence characteristics :( a ) length : 5 amino acids ( b ) type : amino acid ( c ) strandedness : single ( d ) topology : linear ( ii ) molecule type : peptide ( xi ) sequence description : seq id no : 1 : glulyshissergly15 ( 2 ) information for seq id no : 2 :( i ) sequence characteristics :( a ) length : 8 amino acids ( b ) type : amino acid ( c ) strandedness : single ( d ) topology : linear ( ii ) molecule type : peptide ( xi ) sequence description : seq id no : 2 : glulyshisserglyaspilehis15 ( 2 ) information for seq id no : 3 :( i ) sequence characteristics :( a ) length : 15 amino acids ( b ) type : amino acid ( c ) strandedness : single ( d ) topology : linear ( ii ) molecule type : peptide ( xi ) sequence description : seq id no : 3 : glulyshisserglyaspilehisglyalathrtyrserasplys151015 ( 2 ) information for seq id no : 4 :( i ) sequence characteristics :( a ) length : 19 amino acids ( b ) type : amino acid ( c ) strandedness : single ( d ) topology : linear ( ii ) molecule type : peptide ( xi ) sequence description : seq id no : 4 : aspalaasnaspileglualaasnthrtyrasntyrglyglnxaaphe151015asnglugly ( 2 ) information for seq id no : 5 :( i ) sequence characteristics :( a ) length : 11 amino acids ( b ) type : amino acid ( c ) strandedness : single ( d ) topology : linear ( ii ) molecule type : peptide ( xi ) sequence description : seq id no : 5 : alaleuasnalagluasnglnvalargilelys1510 ( 2 ) information for seq id no : 6 :( i ) sequence characteristics :( a ) length : 15 amino acids ( b ) type : amino acid ( c ) strandedness : single ( d ) topology : linear ( ii ) molecule type : peptide ( xi ) sequence description : seq id no : 6 : glulyshisserglyaspilehisglyalathrtyrserasplys151015__________________________________________________________________________