If cell wall hydrolytic enzymes are involv-ed in extension growth, a correlation may be expected between hydrolytic activity of the cell walls and growth rate of the tissue from which the walls are prepared. Epicotyl sections from 0 to 5 mm, 6 to 10 mm, and 11 to 13 mm below the apical hook of pea seedlings (Pisum sativum var. Alaska) have relative growth rates of 100:15:2, respectivelv. The relative fl-glucosidase activities (units/mg wall) Data from several laboratories suggest that glycosidases play a role in wall plasticization, thus permitting cell elongation during extension growth (9). There is evidence that oligosaccharidehydrolyzing enzymes are localized in the walls (1,2,3,7,13,17,19,25) and that these enzymes can hydrolyze cell wall oligosaccharides (12,13, 15,18,25). It becomes important to know whether walllocalized glycosidase activity is correlated with the growth rate of the tissue from which the walls are prepared. Previous studies, mainly employing enzymes extracted from wall preparations (7,13,19,25), report that the amount of glycosidase activity correlates with the growth rate of the tissue from which the walls are prepared. In this current study, highly purified cell walls from Pisum epicotyls were utilized, thus permitting specific activity determinations. We find that (3-glucosidase specific activity is highest in walls from rapidly elongating tissue but that surface sterilized in 1 % sodium hypochlorite for 15 min, soaked in running tap water for 18 hr, and germinated on absorbent paper in covered plastic trays. Germination was in the dark at 25 C and 85c% relative humidity, and seedlings were harvested 84 hr after germination was begun. Growth rates of the seedlings were determined by marking at 2.5-mm intervals and subsequently measuring the spacings. Sequential epicotyl sections were taken from 0 to 5 mm, 6 to 10 mm, and 11 to 15 mm below the apical hook. The sections were either dropped into beakers on Dry Ice immediately after cutting in the case of samples for cell wall preparation by the nonaqueous method, or, in all other cases, they were weighed out in 0.5-g lots and then frozen on Dry Ice as soon as 0.5 g was obtained. The tissue was stored at -80 C until used.Cell Wall Preparation. Cell walls were prepared by homogenization in glycerol and filtration through a glass bead filter, as described by Kivilaan et al. (16). The wall material was dried in an evacuated dessicator over P205, CaCl2, and paraffin at 4 C for 24 to 48 hr. The resultant white powder was stored at -10 C in a sealed jar over CaSO4. Alternatively glycerol was removed from the pelleted walls by water, rather than by solvent washing, as in the Kivilaan procedure. Washing and collection by centrifugation was repeated three times, the cell wall material was collected by filtration, dried over P205 in vacuo, and stored as described above.Aqueous Cell Wall Preparation. Cell walls were also prepared by homogenization of 0.5-g aliquots of frozen tissue sections ground at 0 to 4 C in a 50-ml conical glass hom...

In studies on the degradation of glycogen by rhGAA, a glycosylated protein core material was found which consists of about 5–6% of the total starting glycogen. There was an additional 25% of the glycogen unaccounted for based on glucose released. After incubation of glycogen with rhGAA until no more glucose was released, no other carbohydrate was detected on HPAEC-PAD. Several oligosaccharides are then detectable if the medium is first boiled in 0.1 N HCl or incubated with trypsin. It is present in serum either in an HCl extract or in a trypsin digest. The characteristics of the in vivo serum material are identical to the material in the in vitro incubation medium. One oligosaccharide cannot be further degraded by rhGAA, from the incubation medium as well as from serum co-elute on HPAEC-PAD. Several masked oligosaccharides in serum contain m-inositol, e-inositol, and sorbitol as the major carbohydrates. The presence of this glycosylated protein in serum is a fraction of glycogen that is degraded outside the lysosome and the cell. The glycosylated protein in the serum is not present in the serum of Pompe mice not on ERT, but it is present in the serum of Pompe disease patients who are on ERT, so it is a biomarker of GAA degradation of lysosomal glycogen.

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Antibodies directed against recombinant erythropoietin have been obtained by immunization of rabbits with the hormone in Freund's complete adjuvant. Two sets of antibodies are present in the serum of the immunized rabbits. The results of oxidation of the erythropoietin with periodate, inhibition of the antibodies with the structural monosaccharide residues of the hormone, and reaction of the antibodies with lectins of known carbohydrate specificity have established the antibodies to be anti-carbohydrate antibodies. These antibodies may be of value as tracking agents for some diseases and should be useful for detecting abuses of the hormone in enhancing performance in athletic competitions.