Total Protein Production Research Articles

Equine motor neuron disease: findings in 28 horses and proposal of a pathophysiological mechanism for the disease

Over a three and one-half year period, 28 adult horses were diagnosed with equine motor neuron disease (EMND). The most commonly identified environmental risk factors for a horse having EMND were absence of grazing for more than a year and provision of poor quality hay. Quarter Horses were 5.4 times more at risk than other breeds but this was thought to be an epiphenomenon related to the frequency of Quarter Horses at boarding stables. Weight loss, excessive recumbency and/or trembling were the first signs noted. Other clinical diagnostic signs included: constant shifting of the weight in the rear limbs, abnormally low head carriage and muscle fasciculations. Excellent to ravenous appetites were present in all cases and marked coprophagia in some cases. Abnormally high serum concentration of muscle-derived enzymes was the only consistent serum chemistry abnormality found. Abnormal glucose absorption, increased cerebrospinal fluid total protein and intrathecal production of IgG were identified in a number of cases. Euthanasia was performed on 5 horses within 4 days of hospital admission, because of inability to stand or respiratory distress, and on 18 horses after the diagnosis had been completed. Five affected horses were maintained for observational purposes for periods of 9 months to over 2 years after the onset of clinical signs. They were given access to pasture and 2 were given supplemental vitamin E as the only therapy. Marked clinical improvement occurred in the 4 more acutely affected horses. Pathological findings, preference of type 1 muscle fibre atrophy and lipopigment accumulation within the capillary endothelium of the spinal cord of all cases, supported the hypothesis of EMND being an oxidative disease.(ABSTRACT TRUNCATED AT 250 WORDS)

Dexamethasone potentiates the stimulatory effect of insulin-like growth factor-I on collagen production in cultured human fibroblasts.

We examined the effects of insulin-like growth factor-I (IGF-I) and dexamethasone on the production of collagen by cultures of human infant foreskin fibroblasts, and the interaction between these two factors. IGF-I at 500 ng/ml maximally increased collagen accumulation fourfold. Collagen was increased twofold relative to total protein production. Dexamethasone at a concentration of 1 mumol/l reduced collagen production by between 25% and 40% in unstimulated cells and those cultured with up to 100 ng IGF-I/ml. However, dexamethasone did potentiate collagen production in cells stimulated with 250 ng IGF-I/ml. This potentiation was independent of any effects of IGF-I or dexamethasone on prostaglandin (PG)E2 production. Transforming growth factor-beta (TGF-beta) is also a potent stimulator of collagen formation. However, no potentiation of TGF-beta-stimulated collagen production by dexamethasone was apparent. The mechanism by which dexamethasone potentiates IGF-I-stimulated collagen production was investigated. Dexamethasone treatment increased IGF-I binding to the type 1 IGF receptor without altering the binding affinity. Dexamethasone also attenuated the secretion of IGF-binding proteins by IGF-I-maintained cells.

Glucocorticoid-induced activation of latent transforming growth factor-beta by normal human osteoblast-like cells.

Transforming growth factor-beta (TGF beta) is a multifunctional growth factor in bone that is secreted as a latent complex and must be activated in order to influence cellular activity. We have investigated the influence of dexamethasone (Dex; a potent glucocorticoid) on the secretion and activation of latent TGF beta by normal human osteoblast-like cells (hOBs). Dex had no significant effect on TGF beta mRNA or total protein production, but treatment with Dex resulted in a steroid dose-dependent activation of up to 90% of the TGF beta produced by the hOBs. Dex-treated hOBs activated multiple latent forms of TGF beta. Conditioned medium from Dex-treated hOBs retained the ability to activate latent TGF beta when incubated at 37 C in the absence of cells. Unlike other cell systems, Dex-induced hOB-mediated TGF beta activation did not involve binding to the mannose-6-phosphate receptor. However, the activation was prevented by treatment of hOB cells with microtubual disrupting agents, by the addition of protease inhibitors, or by weak base treatment of the medium. Dex treatment of hOBs was shown to induce a dose-dependent increase in the mRNA levels of cathepsin-B and -D and in the levels of cathepsin-B protein secreted by the cells. Taken together, these data suggest that Dex treatment of hOBs induces the production and secretion of lysosomal proteases that, when secreted, activate latent TGF beta which is secreted by the hOB cells. There is evidence for an involvement of more than one type of protease in this activation process. This activation of TGF beta may, therefore, play a role in glucocorticoid regulation of bone cell functions. Furthermore, TGF beta is most likely involved in autocrine and paracrine effects on bone cells.

Effects of two levels of concentrate supplementation on milk production of dairy goats browsing Mediterranean scrubland

Effects of two levels of supplementation (low (LC) 750 g/d and high (HC) 1500 g/d from d 30 post-partum) on 4 months lactation of 17 Anglo-Nubian × Damascus goats were evaluated. Goats browsed 5–6 h daily on typical Mediterranean scrubland, dominated by Common oak ( Quercus calliprinos) and shrubs (mainly Pistacia lentiscus, Sarcopoterium spinosum and Calicotome villosa), young Cupressus sempervirens trees but few grasses. Goats were separated from their kids 2 d postpartum. HC feeding depressed net grazing time at pasture (75.1% and 59.2%, P<0.05). Although goats in the HC group produced more milk (2.23 vs. 1.81 kg/d, P< 0.05), no significant differences were found in total production of fat, protein and solids for cheese manufacturing (SCM, 188 and 158 g/d, P=0.22). On d 38, milk yield of LC goats was lower (2.31 vs. 2.84 kg/d, P<0.05), and protein (5.54% vs. 4.74%) and fat (3.69% vs. 3.40%) contents were higher, compared to HC ( P<0.05). No differences in milk composition between groups were detected later. However, from d 85 of lactation onward, LC goats yielded 26% less kg milk, 24% less kg protein, 25% less kg fat and 24% less kg SCM ( P<0.10) than HC; LC goats produced less lactose throughout the experiment, but not significantly at d 51 ( P<0.05). Plasma NEFA concentration was higher in the LC than in the HC group at d 38, 85 and 113. Energy balance, as calculated from plasma NEFA values, was negative in the LC goats in all periods but positive in HC from d 85 onward. Since no significant difference in SCM throughout lactation was found, and the main effect of HC feeding was an extra production of lactose, decision to provide HC feeding may be economically justified only if the extra income from lactose (as part of whole milk, yoghurt, or whey-cheese) exceeds the cost of 750 g concentrates.

Crude protein and yield components of six kenaf cultivars as affected by crop maturity

Information concerning the influence of plant maturity on kenaf ( Hibiscus cannabinus L. Malvaceae) as a livestock feed is limited. Research was conducted during 1989 and 1990 to determine the effect of plant maturity on crude protein and yield components of kenaf. Crude protein and yield data were collected on six kenaf cultivars (‘Guatemala 4’, ‘Guatemala 45’, ‘Guatemala 48’, ‘Guatemala 51’, ‘Cuba 2032’ and ‘Everglades 41’) harvested at 76 and 99 days after planting (DAP) at Ladonia, TX, on a Tinn clay, 0–2% slope. Cultivars, harvest dates and years each significantly affected yield components. Crude protein percentages in the leaves, stalks and whole plants were significantly greater at 76 DAP than 99 DAP, while the reverse was true for total crude protein production. Guatemala 48 produced the greatest crude protein (558 kg/ha) of the tested cultivars. The effects of harvest maturity were consistent across cultivars and years, even though differences existed among cultivars for yield components and crude protein. The selections of harvest maturity and cultivar are important variables in producing maximum protein and dry-matter production.

Effect of salmon calcitonin on human adherent monocytes after exposure to heat-shock

We examined the effect of salmon calcitonin on human monocytes after exposure to heat-shock (43 °C for 45 minutes). Trypan blue dye exclusion test showed that monocytes functionally activated by bacterial lipopolysaccharide (LPS) offer a significant survival advantage when exposed to hyperthermia whereas calcitonin-treated phagocytes had a biphasic behavior with a decrease in cell viability 60 minutes after heat-shock. Adherence to plastic increased the living cell number in the monocytes as compared with the cells cultivated in polypropylene tubes. Total protein production analysis indicated that calcitonin blocks macromolecular protein synthesis induced by LPS in both normal and stress conditions, but this synthesis is always increased in calcitonin-treated cells in comparison with the controls. These data indicate that calcitonin is efficiently active in myelomonocytic cells at both physiologic and stress temperatures, increasing cell survival and triggering cell death and suggesting that in monocytes, after heat-shock, a “ready to go” death program is kept in check by molecules such as heat-shock proteins.

Enhanced Production of Morbillivirus Gene-Specific RNAs Following Induction of the Cellular Stress Response in Stable Persistent Infection

Previous in vitro work demonstrated the incorporation of the major inducible 70k heat shock protein (i.e., 72k HSP) into the biologically active light nucleocapsid (L-NC) variant of canine distemper virus (CDV). Here, in vitro induction of the cellular stress response, characterized by elevated cytoplasmic and intranuclear 72k HSP, enhanced L-NC expression in mink lung cells supporting stable persistent infection by raccoon-origin CDV. Increases in L-NC were correlated to increased viral RNA production in cell-free transcriptional assays. The enhanced production of viral transcripts within infected cells following stress response induction was confirmed by slot blot and Northern blot analysis of total cellular RNA and was reflected in increased total viral protein production. Post-shock increases in viral fusion (F) gene transcripts and F protein were associated with dramatic increases in viral cytopathic effect. Modest induction of cell-free infectious viral progeny was also documented. A similar effect of the cellular stress response upon vital protein expression, cytopathic effect, and cell-free infectious progeny release was demonstrated in murine neuroblastoma cells persistently infected with a canine CDV isolate. Alterations of the persistent viral phenotype were independent of the specific mechanism of stress-response induction (i.e., heat or sodium arsonite), supporting the role of the stress response and not a particular stressor in mediating these changes. These results document the ability of the cellular environment to alter persistent viral RNA metabolism, thereby altering the infection phenotype.

Role of interleukin-6 in mediating the acute phase protein response and potential as an early means of severity assessment in acute pancreatitis.

A number of laboratory and clinical studies have shown that interleukin-6 is the principal mediator of the acute phase protein response. In this study the relationship between serum concentrations of interleukin-6 and C-reactive protein in acute pancreatitis are examined and the ability of interleukin-6 to discriminate between severe and mild attacks is assessed. We have studied 24 patients (10 severe and 14 mild). Serum samples were collected on admission, six hourly for 48 hours and then 12 hourly for a further three days. When the areas under the curves of individual patients were compared there was a strong correlation between the total production of interleukin-6 and C-reactive protein (r = 0.73) (Spearman rank correlation) and peak interleukin-6 and C-reactive protein concentrations (r = 0.75), suggesting a close relationship between interleukin-6 and C-reactive protein production. Both on admission and peak interleukin-6 concentrations were significantly higher in patients with severe than mild disease. There was no significant difference in on admission C-reactive protein concentrations, although significant differences were seen when peak concentrations were considered. Utilising a peak interleukin-6 concentration of > 130 u/ml, we were able to distinguish between severe and mild attacks of acute pancreatitis with a sensitivity of 100% and specificity of 71%. These figures were comparable with those for peak C-reactive protein, a C-reactive protein of > 150 mg/l detecting severe attacks of acute pancreatitis with a sensitivity of 90% and specificity of 79%. In view of the fact that interleukin-6 concentrations peaked earlier than those of C-reactive protein, interleukin-6 is capable of providing comparable, but earlier severity prediction than C-reactive protein.

Macrophage and monocyte IL-1 beta regulation differs at multiple sites. Messenger RNA expression, translation, and post-translational processing.

Maturation of blood monocytes into macrophages is accompanied by a number of functional changes including decreased IL-1 beta release in response to LPS. This limitation has previously been ascribed to transcriptional regulation. However, in seeming conflict with the observed depression in IL-1 beta mRNA levels, recent work demonstrates increased intracellular IL-1 beta in macrophages. Therefore, the present study sought to explain these differences by comparing IL-1 beta production from autologous alveolar macrophage and blood monocyte pairs at multiple regulatory sites, including endotoxin responsiveness, mRNA expression, protein translation, and post-translational processing. Macrophages did not differ from monocytes in endotoxin sensitivity, but when analyzed by both ELISA and Western blot, were confirmed to have limitations in IL-1 beta release. Gene expression studies demonstrated that at 4 h, macrophage IL-1 beta steady state mRNA levels were 3-fold lower than the monocyte's. However, total IL-1 beta protein production, as measured by [35S]methionine labeling with immunoprecipitation, demonstrated three- to sixfold higher amounts in macrophages at comparable time points. The enhanced protein production in the face of relatively low mRNA levels suggests that macrophages translate IL-1 beta mRNA more efficiently. Furthermore, characterization of IL-1 beta release into supernatants revealed that whereas monocyte release occurred early, represented 5 to 20% of the intracellular amounts, and contained largely processed IL-1 beta, macrophage release was delayed, represented 1 to 5% of the intracellular amounts, and contained primarily unprocessed IL-1 beta. Taken together, these data demonstrate that the limitations in alveolar macrophage IL-1 beta release occur due to slower export and conversion of 35- to 17-kDa protein and are not due to differences in sensitivity to endotoxin or to transcriptional control mechanisms.

Failure of tachykinins including substance P and its fragments to influence proteoglycan and protein synthesis in bovine chondrocytes in vitro

Adult bovine articular chondrocytes were exposed to substance P, neurokinins A and B or substance P fragments, SP 1–4, SP 1–6 and SP 7–11 in vitro. Proteoglycan synthesis was assessed by measuring proteoglycans which were released into the culture medium or incorporated into the cell layer. The intact tachykinins or substance P fragments had no direct effect on proteoglycan synthesis. Nor was total protein production affected. Gel chromatography, under dissociative conditions, revealed that sulphated proteoglycans detected in the medium or cell layer following treatment of chondrocytes with substance P, contained proteoglycans of similar molecular weight to those produced by cells exposed only to diluent controls. Therefore, we conclude that the acceleration of arthritis by substance P does not appear to be mediated through an effect on chondrocyte synthetic function.

Effect of SMS 201-995 on exocrine pancreatic secretion in a patient with external pancreatic fistula.

The effect of SMS 201-995 on pancreatic exocrine function was studied. The SMS 201-995 was administered to a patient with an artificial external pancreatic fistula following pancreaticoduodenectomy. Variations in pancreatic exocrine function were assessed by determining the volume and components of the fistula fluid during the following periods: 5 d prior to SMS 201-995 administration, for 5 d during actual administration, and for 5 d after it had been discontinued. The SMS 201-995 was administered by subcutaneous injection of 100 micrograms every 12 h for the first 2 d and then 100 micrograms every 6 h for 3 d. This experiment demonstrated that SMS 201-995 has a strong inhibitory effect on pancreatic exocrine function, markedly reducing the amount of fistula fluid and the production of amylase, total protein, and bicarbonate.

Evaluation of a leukotriene receptor antagonist in prevention of hyperoxic lung injury in newborn rabbits

Prolonged exposure to hyperoxia can result in significant lung injury and has been associated with the development of bronchopulmonary dysplasia. Leukotrienes (LT) recruit polymorphonuclear leukocytes (PMN) to the lung, increase vascular permeability, and have therefore been postulated to play a role in the pathogenesis of hyperoxic lung injury. This study investigates ICI 198,615 (ICI), and LTD 4 and LTE 4 receptor antagonist in preventing hyperoxic lung injury in newborn rabbits. Matched littermates of 7-day-old rabbits received ICI (0.1 or 1.0 μM/kg/h) or vehicle alone, were exposed to >95% O 2, and sacrificed after 48, 72, 84 and 96 h of exposure. Bronchoalveolar lavage fluid (BAL) of the left lung was analyzed for white cell count, differential, absolute number of PMNs, total protein, and cyclooxygenase products 6-keto-PGF 1α, and thromboxane B 2. Lung water was quantified utilizing the right lung. Results demonstrated no significant differences between the ICI groups or between the ICI groups and controls. In conclusion, the administration of the LTD 4 and LTE 4 receptor antagonist ICI 198,615 was insufficient to reduce the formation of pulmonary edema, reduce mortality or attenuate hyperoxic lung injury. These experiments suggest that a number of other mediators may be involved in the hyperoxic lung injury process and that the functional inhibition of a portion of the arachidonic acid cascade was not sufficient to either prevent or attenuate hyperoxic lung injury in newborn rabbits.

Rhizopus oligosporus grown on natural rubber waste serum for production of single cell protein: a preliminary study

Maximum production of mycelium and utilization of total organic carbon byR. oligosporus grown on natural rubber waste serum was achieved at 28°C with an inoculum size of 7.5% (v/v) and grown for 144 h with an initial pH of 4.0. The maximum production of total crude protein, however, was when culture medium was inoculated with 2% (v/v) of spore suspension under the same conditions. Natural rubber waste serum may be a potential substrate for the production of single cell protein.

Inter-relationships among total crude protein and digestible dry matter production and their component traits in Guinea grass

Phenotypic and genotypic correlation coefficients of total crude protein per plant (TCP) and digestible dry matter per plant (DDM) with dry matter yield per plant (DMY), tiller number, plant height, stem thickness and leaf-stem ratio, pooled over two years, revealed that TCP and DDM were correlated at both the phenotypic and genotypic levels with DMY, tiller number and plant height. But only DMY had high direct effects on TCP and DDM. Tiller number and plant height had only indirect effects on TCP and DDM via DMY. Thus, selection based on tiller number and plant height may not only improve dry matter yield per plant but also total crude protein per plant and digestible dry matter per plant. Further, screening of high TCP and high DDM clones among the high yielding clones would be a more remunerative approach to improve TCP and DDM in Guinea grass.

Characteristics of batch culture of a recombinantbacillus brevis excreting a foreign esterase

The productivity of extracellular enzyme was evaluated in batch culture using a protein hyperexcreting host, Bacillus brevis HPD31(5) harboring pHSC131, which carried a gene (est) encoding esterase activity from Bacillus stearother mophilus. Optimum temperature and pH for the bacterial growth and the production of extracellular esterase were found to be 35 degrees C and pH 6.5, by using the standard medium (GPY) containing neomycin as a selective pressure, Under the cultivation condition employed, cell growth reached 5 g dry cell weight/L, while the extracellular esterase activity amounted to 4.5 U/mL. Most (79%-92%) of the esterase produced was excreted into the medium. pHSC131 was stably retained in the host cell during cultivation in the presence of neomycin. However, in the absence of neomycin, the plasmid was completely lost from the host after 12-h cultivation accompanied by decreases in both esterase activity and production of total extracellular protein. The copy number of the plasmid was estimated to be approximately 7 throughout the cultivation. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the excreted proteins showed the presence of a protein having an apparent molecular weight of 32,000, which equals to the value predicted from the DNA sequence of the est gene.

The effects of chronic exposure to supraphysiological concentrations of 3,5,3? triiodo-L-thyronine (T3) on cultured GC cells

We have characterized the effects of supraphysiological concentrations of T3 on GC cells, a cultured cell line in which physiologic concentrations of T3 regulate cell growth, protein content, and growth hormone (GH) production. GC cells were exposed to 3 times (1.0 nM) and 80 times (25.0 nM) the physiologic concentration of T3 (0.3 nM) for either 4 d or for greater than 3 months. Both short and prolonged exposure to supranormal T3 concentrations supported maximal cell growth rate and induced significant increases in total protein (P less than 0.025) and GH production (P less than 0.01) per cell when compared to measurements in control GC cells. In addition, exposure to 1.0 nM and 25.0 nM T3 for greater than 3 months enhanced the toxicity of heat shock in a manner similar to previously described effects on GC cells due to T3 exposure of shorter duration. Thus, initial responses to raised T3 concentrations in cultured GC cells persisted without alteration when hormone exposure was prolonged for greater than 3 months.

Inhibition of Collagen Production by Diabetic Rat Serum: Response to Insulin and Insulin-Like Growth Factor-I Addedin Vitro*

Diabetes mellitus is associated with a generalized defect in connective tissue metabolism. Since collagen is the major protein of connective tissues, we used collagen as a probe to examine the role of factors in diabetic rat serum (DRS) in the etiology of these defects. Serum and skin fibroblasts were isolated from nondiabetic rats, and serum was taken from rats 48 h after injection of 200 mg/kg streptozotocin. Within 24 h of confluency, the fibroblast medium was changed to experimental serum for 24 h, with 5 microCi [3H]proline added for the final 6 h. Collagen and noncollagen proteins were quantitated using purified collagenase. Compared to cells incubated in medium without serum, collagen fell to 58% with 0.5% DRS (P less than 0.05) and continued to decrease with increasing concentrations of DRS. Noncollagen protein decreased below levels in cells incubated in medium without serum only when concentrations of diabetic serum were 1% or greater and did not decrease further with higher concentrations of diabetic serum. Collagen was decreased to a greater degree than noncollagen protein at each concentration of DRS, such that collagen relative to total protein production was significantly reduced at 0.5% or more DRS. Addition of 10(-7)-10(-9) M insulin or insulin-like growth factor-I (0.1-1000 ng/ml) to DRS did not return collagen production to the level seen in cells incubated in medium with no added serum (basal production). After separation of serum components based on size, incubation of cells with the low mol wt fraction (less than 5000 daltons) of normal and diabetic rat serum resulted in equivalent collagen production, while incubation with the high mol wt fraction of DRS resulted in 200-fold less collagen compared to the similar fraction of normal serum. This decrease in collagen production appeared due to the presence of a high mol wt factor(s) in diabetic serum which had a direct inhibitory effect on collagen and was not due to deficiency of growth peptides. The degree and specificity of these changes in collagen production probably contribute to long term complications in diabetes through altered connective tissue metabolism.

Stimulation of Collagen and Glycosaminoglycan Production in Cultured Human Adult Dermal Fibroblasts by Recombinant Human Interleukin 6

Interleukin (IL) 6 is a pleiotropic cytokine synthesized by fibroblasts in response to many stimuli, including IL-1 beta. To evaluate the possibility that previously observed stimulation of fibroblast biosynthetic functions by IL-1 beta may be mediated by autocrine IL-6, we investigated the effect of recombinant human (rh) IL-6 on the connective tissue-related biosynthetic functions of three lines of cultured human adult dermal fibroblasts. We found that rhIL-6 mimicked some of the activities of IL-1 beta, as 24-96-h treatment of confluent fibroblast cultures with rhIL-6 caused concentration (10 to 1000 ng/ml)-dependent increases in the production of collagen and the glycosaminoglycans (GAG), hyaluronic acid and chondroitin-4/6-sulfates, but had little effect on fibronectin or total protein production. Although the effective stimulating concentrations of IL-6 were within the range (approximately 100 ng/ml) we found produced by rhIL-1 beta-treated fibroblast cultures, rhIL-1 beta at 0.2-1.0 ng/ml induced significantly greater amounts of collagen and GAG than the maximum effective concentrations of IL-6. Moreover, an anti-rhIL-6 antibody, which effectively neutralized the fibroblast-stimulating activities of rhIL-6, only fractionally blocked the fibroblast-stimulating actions of rhIL-1 beta, suggesting autocrine IL-6 only partially mediates the effects of IL-1 beta on fibroblasts. Conversely, the fibroblast-stimulating effects of rhIL-6 are unlikely due to autocrine IL-1 beta, as an anti-rhIL-1 beta antibody had only minimal inhibitory action on rhIL-6-treated fibroblast cultures. Overall these results suggest that IL-6 could function as a paracrine/autocrine regulator of dermal fibrotic repair.

The in vitro effects of metal cations on eukaryotic cell metabolism

The in vitro cytotoxicity of nine metal cations common in dental casting alloys was evaluated using Balb/c 3T3 fibroblasts and four toxicity parameters: total protein production, 3H-leucine incorporation, 3H-thymidine incorporation, and MTT-formazan production. Concentrations causing 50% toxicity compared to controls (TC50's) and reversibility of these effects were determined. The range of potency of the metal cations was 2-3 orders of magnitude, with Cd2+ showing the greatest potency and In3+ showing the least. Potency did not correlate with atomic weight for these metals. For each metal cation, the TC50's of the various toxicity parameters were similar in most cases. However, several cations (Cu2+, Ga3+) showed greater potency with 3H-thymidine incorporation. Reversibility of the toxic effects was observed for all cations; the effects generally became irreversible at concentrations in the range of the TC50 value for each cation. Several stimulatory effects were seen. Small but statistically significant stimulations were observed after 24 h of metal exposure for Ag1+, Au4+, Cu2+, Ga3+, and Ni2+. Residual stimulations 24 h after removal of the metal cations were observed for Au4+, Cd2+, Ni2+, and Zn2+. Stimulations always occurred at concentrations below the TC50 concentrations. This study should be useful in evaluating the potential cytotoxic effects of metal cations released from dental alloys.

Biotechnology of Seed Crops: Genetic Engineering of Seed Storage Proteins

Table 1. Amino acid composition of total maize and soybean seed protein and purified storage protein fractions.z During their development, seeds accumulate large amounts of storage, or reserve, products that are metabolized during germination and the early stages of seedling growth. Energy reserves include complex carbohydrates (starch) and lipids, while reserves of N, C, and S are accumulated in specific storage proteins. The uses of seed products by man are innumerable, ranging from fermentation of maize starch for alcohol production to refinement of soybean protein for use in infant formula. Agronomic crops, such as maize, wheat, oats, and soybeans, are grown because they generate large amounts of seed reserves per unit of cultivated land. A major concern of the agricultural industry is increasing yields of seed components to enhance the value of these crops. During the past century, plant breeders have been quite successful in developing new cultivars of seed crop plants that give higher yields of seed components than older ones. Advances have been made not only with respect to increasing the content of starch, oil, and protein in the seed, but also, in some cases, in altering the quality of these components. For example, naturally occurring genetic variation has been exploited in the development of maize cultivars with modified starch content (Nelson, 1979; Shannon and Garwood, 1984) and increased or altered oil content (Alexander 1988, 1989; Weber, 1987). In a similar manner, the composition of soybean oil has been altered through selection to decrease the content of linolenic acid, resulting in a product with a more desirable flavor (Fehr, 1989; Hymowitz, 1987). The synthesis of carbohydrates and lipids involve complex metabolic processes, and although a great deal is known about these pathways, it remains to be determined how recent advances in genetic engineering technology will aid in the modification of these processes to enhance seed quality. For example, alteration of the activities of one or two enzymes in the starch biosynthetic pathway might allow for the accumulation of desirable intermediates of starch biosynthesis. Such a situation may, however, result in a serious imbalance in the overall carbon flow in the seed, which could affect factors such as grain yield and seed viability. Conventional selection and breeding strategies may, therefore, be more efficient than the use of genetic engineering in the development of crop cultivars possessing seed carbohydrates and lipids of desirable composition (Alexander, 1988). An important aspect of seed quality that is directly amenable to manipulation through genetic engineering involves the nutritional characteristics of seed proteins. In most cases, seed proteins of any one crop do not contain sufficient quantities of certain amino acids that are essential in the diet of humans and other monogastric animals (Nelson, 1969). Generally, cereals are most limiting in lysine and tryptophan, and legumes are deficient in the sulfur amino acids methionine and cysteine (Table 1). The economic advantages for increasing the nutritional quality of seed proteins are enormous: estimates of total protein production for each of two major seed crops in the United States, maize and soybeans, exceed 16 million tons annually (Wilson, 1987). Development of nutritionally balanced seed proteins in high-production crops like maize and soybean would benefit all areas of the agricultural industry concerned with grain production for human and livestock consumption. Plant breeders have had only limited success in improving the nutritional quality of seed proteins, primarily because genes encod-