Light-chain framework region residue Tyr71 of chimeric B72.3 antibody plays an important role in influencing the TAG72 antigen binding.

The crystallographic study of chimeric B72.3 antibody illustrated that there are three FR side-chain interactions with either CDR residue's side chain or main chain. For example, hydrogen bonds are formed between the hydroxyl group of threonine at L5 in FR1 and the guanidinal nitrogen group of arginine at L24 in CDR1, between the hydroxyl group of tyrosine at L36 in FR2 and the amide nitrogen group of glutamine at L89 in CDR3 and between the hydroxyl group of tyrosine at L71 in FR3 and the carbonyl group of isoleucine at L29 as well as the amide nitrogen group of serine at L31 in CDR1. Elimination of these hydrogen bonds at these FR positions may affect CDR loop conformations. To confirm these assumptions, we altered these FR residues by site-directed mutagenesis and determined binding affinities of these mutant chimeric antibodies for the TAG72 antigen. We found that the substitution of tyrosine by phenylalanine at L71, altering main-chain hydrogen bonds, significantly reduced the binding affinity for the TAG72 antigen by 23-fold, whereas the substitution of threonine and tyrosine by alanine and phenylalanine at L5 and L36, eliminating hydrogen bonds to side-chain atoms, did not affect the binding affinity for the TAG72 antigen. Our results indicate that the light-chain FR residue tyrosine at L71 of chimeric B72.3 antibody plays an important role in influencing the TAG72 antigen binding. Our results will thus be of importance when the humanized B72.3 antibody is constructed, since this important mouse FR residue tyrosine at L71 must be maintained.     

Factors affecting central inspiratory modulation of hypoglossal motoneuron activity in newborn pigs

In newborn pigs (4-6 days old), recordings of efferent whole hypoglossal and phrenic nerve discharges were obtained during hyperoxia (or normoxia) and during hypoxia, before and after bilateral vagotomy. With intact vagi, spontaneous hypoglossal inspiratory activity was not observed and was not elicited by either spontaneous changes of electroencephalogram (EEG) or hypoxic stimulation (15% O2 in N2). After bilateral vagotomy, some animals had episodes of spontaneous hypoglossal inspiratory activity; power spectral analysis of EEG demonstrated that this inspiratory activity appeared synchronously with shifts of major peaks in EEG spectra from the delta band (0.5-3.5 Hz) to the theta band (3.5-7.0 Hz). Hypoglossal inspiratory discharges were also elicited by hypoxic stimulation and usually had a decrementing discharge pattern; in some cases, this activity had an augmenting discharge pattern. Our results suggest that hypoglossal motoneurons are poorly modulated by central inspiratory drive, requiring additional facilitatory influences, i.e. corticobulbar, intra-bulbar, chemical drive, before such modulation is observed.     

Changes in fructose & citric acid in accessory glands of reproduction of rats following long-term treatment with isomers of clomiphene citrate

The effects of 500 mcg/kg/day of cis- or trans-clomiphene for 60 days on the testis and accessory glands of reproduction in adult male rats and the pattern of recovery following withdrawal of treatment were described. In animals treated with cis-clomiphene, testis weights showed little change from those of controls except in Week 2 of recovery when testicular weights were significantly less than in controls (p.001). The weights of all the accessory glands were significantly less as compared with controls (p.001) at the termination of treatment but reached control levels by the end of the 4 week recovery period. Citric acid (seminal vesicles and ventral prostate) and fructose (dorsolateral prostate and coagulating glands) levels decreased significantly in Week 2 compared with controls (p.001) but were restored to normal by Week 4. Epididymal spermatozoa were nonmotile at the end of treatment as well as in Week 1 of recovery; none of the treated males mated. Spermatozoa motility and normal mating behavior were restored by Weeks 3 and 4 of recovery. In rats treated with trans-clomiphene testes and accessory glands and their secretory activity (citric acid and frutose) were reduced significantly (p.001) and returned to control levels by Week 4 of recovery. Epididymal spermatozoa were nonmotile at the end of treatment and during the first 2 weeks of recovery but returned to normal during Week 3 of recovery along with normal mating behavior. The observed inhibitory effects on the testes and accessory glands may be due to estrogenicity of the compounds modifying the synthesis and/or release of gonadotropins mediated through the hypothalamo-hypophyseal axis. Complete recover, as evidenced by birth of normal young within 30 days of treatment withdrawal, suggests that the effects of these compounds are transient in adult male rats. The trans-isomer seemed to be more estrogenic than the cis-isomer.     

Chronic Alcohol Consumption Increases Cyclo (His-Pro)-Like Immunoreactivity in the Rat Brain

Abstract: Administration of histidyl-proline diketopiperazine (cyclo (HisPro)) to rats attenuates ethanol-induced sleep. To understand the role played by cyclo (His-Pro) in the pathophysiology of prolonged alcohol consumption, we have measured the distribution of this peptide in brains of control and alcohol-treated rats. The data show that prolonged alcohol consumption increases the concentration of cyclo (His-Pro) in hypothalamic as well as extrahypothalamic brain. These changes may reflect a physiologic adaptation of the brain during alcohol consumption.     

Varietal response to iron chlorosis in upland rice

Summary In a study involving twenty upland rice genotypes, the induction of iron chlorosis was found to be more by superphosphate application than due to lime incorporation into an alfisol soil under greenhouse conditions in GI tray-grown seedlings as quantified by two parameters viz., total chlorphyll content and Fe²⁺ content. Of the two indices of iron chlorosis, Fe²⁺ was more sensitive than chlorophyll content. Genotypes were grouped into efficient and inefficient categories both in terms of absorption and utilization of iron based on the degree of reduction in response to added superphosphate.     

Effect of phospholipase A on the structure and functions of membrane vesicles from Mycobacterium phlei.

The phospholipid composition of the electron transport particles and coupling factor-depleted electron transport particles of Mycobacterium phlei are the same, but they differ in contents. The accessibility of partially purified phospholipase A to these membrane phospholipids was found to be different. Treatment of membranes of Mycobacterium phlei with phospholipase A impairs the rate of oxidation as well as phosphorylation. The inhibition of phosphorylation can be reversed by washing the membranes with defatted bovine serum albumin. The reconstitution of membrane-bound coupling factor-latent ATPase activity to phospholipase A-treated depleted electron transport particles and their capacity to couple phosphorylation to oxidation of substrates remained unaffected after phospholipase A treatment. However, the pH gradient as measured by bromthymol blue was not restored after reconstitution of phospholipase A-treated depleted electron transport particles with membrane-bound coupling factor-latent ATPase. These findings show that the phosphorylation coupled to the oxidation of substrates can take place without a pronounced pH gradient in these membrane vesicles. The dye 1-anilino-8-naphthalene sulfonic acid (ANS) exhibited low levels of energized and nonenergized fluorescence in phospholipase A-treated membranes. This decrease in the level of ANS fluorescence in phospholipase A-treated membranes was found to be directly related to the amount of phospholipids cleaved. The decrease in the energy-dependent ANS response in phospholipase A-treated electron transport particles, as compared with untreated electron transport particles, was shown to be a result of a change in the apparent K-d of the dye-membrane complex, and of a decrease in the number of irreversible or slowly reversible binding sites, with no change in the relative quantum efficiency of the dye. The decrease in ANS fluorescence in phospholipase A-treated particles appears to be due to a decrease in the hydrophobicity of the membranes.     

Cell attachment and chemotaxis can utilize the same peptide sequence of fibronectin

Fibronectin and fragments of the parent molecule are involved in cell-substrate attachment, cell migration and chemotaxis. The cell attachment sequence, Gly-Arg-Gly-Asp-Ser-Pro, was synthesized and tested for its ability to effect ligamentum nuchae fibroblast chemotaxis. This hexapeptide and fibronectin itself both caused directed cell migration, with maximal activity in the 10(-10) to 10(-9) M range. These data demonstrate that the fibronectin cell binding site and chemotaxis site share a common sequence.