A new cnzymc ctltalyzing the dcamidation of seed sloragc proteins was found in gcrminaling wheat grains and was partially purified. It also acts on egg lysozymc. horse hemoglobin and reduced RNAsc, glutuminc and Gly-L-Gin-t.-Tyr, No activity was obscrvcd when using ovnlbumin, serum albumin, RNAsc, insulin, asparagine and an asFaraginc_contailling pcptidc. Only glutaminyl rcsiducs appar to bc dcamidated by this cnryme.It differs from trunsglulaminasc and proved to be a true protein dcdmidasc.Protein dcamidilsc; Germinating seed: Wheat INTRODUCTSQNThe degradation of storage proteins during seed germinatio~ is a complex multistage process [ 11. Deamidation preceding their complete proteolytic breakdown was observed in germinating seeds of several leguminous species [Z-4] and is especially intensive in germinating wheat grains [S]. Up to 35% of amido groups of gluten -the complex of wheat grains protein -is deamidated during the first 3 days of germination [S]. It may be inferred from this that the process of deamidation of storage protein in germinating seeds is of a general nnture. The high rate of deamid~tion is indicative of ils enzymatic character,The objectives of the present work were the detection, partial purification and characteriation of the deamidating enzyme from gernlinating wheat grains. The latter were chosen as the enzyme source due to the highest content of amidated amino acids in their storage proteins, which is suggestive of a higher activity of the deamidating enzyme. MATERIALS AND METHODS 3.1, Substrms IL'SI~The Following proteins were isolated using the merhods described in the rci'erenccs: isoelectri~lly pr~cipi~Led soybcvn globulins 161, soybean I1 S and 7 S proteins [7] The incubi&on mixlurc consisted of 0.2 ml of subrrrale solution (3 mpml of prouin or 0,s mg/ml ofamino acid or p;ptidc) in phosphalccitrate buffer. pH 6.8 and ionic strenght 0.067, and 0,4 ml of cnzymc solution. In the control ex~rirn~nts ihc substrate solution was subs& tuted for 0.2 ml of the buffer. After 30 min incubation at 30DC the rdaclion was stoaacd bv the addhion of 0.5 ml of 4% solution of ~&,o,~ 10HIO' 'con&ing enough NaOH to bring up the nH or the incubation mixture to IO. NH, was distilled isothcnnallr Into I M i-i ?SO1 and de~crmincd by a v&&L of the Ncsslcr's mcth& fl I]. All the delermi~ations were ~rformed at fcast in t~pli~t~~x~pt in the chrometographic fractions whcrc they were made in duplicate.Soybean globulin was used as a working subslratc during purificalion and study of the properties of the enzyme. Its natural substrate, gluten. is iess suitable for the determination of the activity in vitro bccausc as a result ofclulnps of gluten ai the shinning of incubation, the reaction proceeds slowly and the repl~ucibi~i~y is poor. The enzyme activity was cxprcsscd in mU (nmol of NH> rclLzs& per inin),The content of the amide N in the substrate prior to and afler incubation with cnzymc was determined by measuring NH) rclcased by.hydrolysis with 2 N HCI [I?]. Transgluiaminasc activity was tcslcd using NH?OH ...

Enzymes catalyzing the deamidation of seed storage proteins were found in germinating seeds of kidney beans (Phaseolus vulgaris L. cv. Moldavian) and squash (Cucurbita pepo L. cv. Mozoleevskaya). They were partially purified and characterized. The properties of these enzymes closely resemble those of the protein deamidase in germinating wheat grains. The detection of similar protein deamidases in germinating seeds of plants belonging to phytogenetically unrelated species indicates that protein deamidases are of considerable physiological significance.

Primed in Ueimiark -aii rigttls rsseri'ed Copyrighi © Physiotof^ia Piantaruill 1996 tSSNOnil-9317 Protein deamidases from germinatiog seeds losif Vaintraub, Ludmila Kotova and Ranajid Shaha Vaintraub, L, Kotova, L. and Shaha, R. 1996. Protein deamidases from germinating seeds. -Physiol. Planl. 96: 662-666.Enzymes catalyzing the deamidation of seed storage proteins were found in germinating seeds of kidney beans {Phaseolus vulgaris L. cv. Moldavian) and squash (Cucurbita pepo L. cv. Mozoleevskaya). They were partially purified and characterized. The properties of these enzymes closely resetnble those of the prolein deamidase in germinating wheat grains. The detection of similar protein deamidases in germinating seeds of plants belonging to phylogenetically unrelated species indicates that protein deamidases are of considerable physiological significance.