COMPETITIVE INHIBITION BETWEEN OXYTOCIN AND LUTEINIZING HORMONE-RELEASING FACTOR (LRF) FOR THE SAME ENZYME SYSTEM IN THE RAT HYPOTHALAMUS

1974 ◽  
Vol 75 (3) ◽  
pp. 435-442 ◽  
Author(s):  
E. C. Griffiths ◽  
K. C. Hooper
Keyword(s):  
Luteinizing Hormone ◽  
Competitive Inhibition ◽  
Enzyme System ◽  
Reproductive Function ◽  
Kinetic Studies ◽  
Supernatant Fraction ◽  
Peptidase Activity ◽  
Rat Hypothalamus ◽  
Enzyme Substrate ◽  
Inhibition Studies

ABSTRACT From the observation that hypothalamic peptidases inactivating oxytocin might be involved in luteinizing hormone-releasing factor (LRF) metabolism and that the hypothalamus contains enzymes which can inactivate LRF, competitive inhibition studies were performed to determine if the peptidases inactivating oxytocin could act on LRF. In mixed substrate incubations, LRF significantly decreased peptidase activity in a hypothalamic supernatant fraction whereas thyrotrophin-releasing factor (TRF) had no effect. From kinetic studies, it was found that there was competitive inhibition between the two polypeptides for the same enzymes and that LRF may be true enzyme substrate. The results confirm the previous proposals that the hypothalamic peptidases can inactivate LRF and the possibility of a role for the enzymes in the regulation of reproductive function is suggested.

THE PRESENCE OF PEPTIDASES IN THE RAT HYPOTHALAMUS INACTIVATING LUTEINIZING HORMONE RELEASING HORMONE (LH-RH)

1974 ◽  
Vol 77 (3) ◽  
pp. 435-442 ◽  
Author(s):  
E. C. Griffiths ◽  
K. C. Hooper ◽  
S. L. Jeffcoate ◽  
D. T. Holland
Keyword(s):  
Luteinizing Hormone ◽  
Reproductive Function ◽  
Assay Method ◽  
Supernatant Fraction ◽  
Peptidase Activity ◽  
Female Rat ◽  
Luteinizing Hormone Releasing Hormone ◽  
Releasing Hormone ◽  
Rat Hypothalamus ◽  
Lh Rh

ABSTRACT The presence of peptidases in the rat hypothalamus inactivating luteinizing hormone-releasing hormone (LH-RH) has previously been demonstrated using an indirect assay method. With the development of a sensitive and specific radioimmunoassay for the releasing hormone, this technique was employed in the study of the peptidases inactivating LH-RH. It was found that supernatant fractions from both male and female rat hypothalami rapidly inactivated LH-RH, and that the peptidase activity of the supernatant fraction was higher in male than in female animals though the particulate fraction's activity was about the same in both sexes. Peptidase activity was also considerably greater in the supernatant than in the particulate fraction. These results confirm that the hypothalamus contains peptidases capable of inactivating LH-RH and give a direct indication that the enzymes may be involved in the central nervous system's control of reproductive function.

scholarly journals SIMULTANEOUS HYDROLYSES OF ESTERS AND PROTEINS AT SATURATION LEVELS

1958 ◽  
Vol 41 (3) ◽  
pp. 485-500 ◽  
Author(s):  
M. Castañeda-Agulló ◽  
Luz M. Del Castillo
Keyword(s):  
Enzyme System ◽  
Kinetic Studies ◽  
Direct Titration ◽  
Enzyme Substrate ◽  
Constant Ph ◽  
Potentiometric Measurement ◽  
The Individual ◽  
Hydrolysis Of ◽  
Protein Enzyme

A direct titration method for the determination of proteolytic activity is discussed. This involves the potentiometric measurement of the volume of 0.08 N NaOH required to maintain a constant pH (8.0) during the time of the hydrolysis. It is a sensitive method which presents several advantages; viz., it measures simultaneously protease and esterase activity, it follows the hydrolysis very closely and from the first stages; the titration is continuous and on the same sample. This method determines a constant fraction of the groups titratable by formol titration. The ratio formol: direct titration is represented by a factor "f" which is presumed to be distinct for each protein-enzyme system. Kinetic studies, using this method, revealed that the rates of hydrolysis of mixtures casein-gelatin on one hand, casein-BAEE or gelatin-BAEE on the other, are always larger than those of the corresponding isolated substrates. In many cases the resulting rates are equal or nearly equal to the sum of the individual rates, even though the mentioned rates have been determined within the saturation zones for every substrate. The former observations are inconsistent with the theory of the formation of an intermediary enzyme-substrate compound, unless it is assumed that the enzyme has a specific active group for each substrate.

THE EFFECT OF OVARIECTOMY ON THE ACTIVITY OF CERTAIN ENZYMES IN THE FEMALE RAT HYPOTHALAMUS

1972 ◽  
Vol 69 (2) ◽  
pp. 249-256 ◽  
Author(s):  
E. C. Griffiths ◽  
K. C. Hooper
Keyword(s):  
Enzyme Activity ◽  
Luteinizing Hormone ◽  
Inverse Relationship ◽  
Supernatant Fraction ◽  
Female Rat ◽  
Hormone Release ◽  
Similar Relationship ◽  
Hormone Levels ◽  
Rat Hypothalamus ◽  
Detectable Difference

ABSTRACT Previous work in the rabbit has shown that the activity of certain peptidases in the hypothalamus which inactivate oxytocin, changes with stimuli known to release gonadotrophins, and may be used as an index of gonadotrophin hormone release (Hooper 1966a,b, 1968; Frith & Hooper 1971a,b). Using this approach, a study was made of the activities of similar peptidases in the rat hypothalamus following ovariectomy, a condition known to cause gonadotrophin release. Enzyme activity in the supernatant fraction was found to decrease progressively with time after ovariectomy, until 42 days after operation, thereafter maintaining a level not significantly different from that at 42 days; there was no detectable difference in particulate enzyme activity after ovariectomy. An inverse relationship between supernatant enzyme activity and luteinizing hormone levels is suggested. It is concluded that a similar relationship to that in the rabbit exists between enzyme activity in the rat hypothalamus and the release of luteinizing hormone-releasing factor from the tissue.

THE EFFECTS OF ORCHIDECTOMY AND TESTOSTERONE PROPIONATE INJECTION ON PEPTIDASE ACTIVITY IN THE MALE RAT HYPOTHALAMUS

1973 ◽  
Vol 72 (1) ◽  
pp. 1-8 ◽  
Author(s):  
E. C. Griffiths ◽  
K. C. Hooper
Keyword(s):  
Luteinizing Hormone ◽  
Testosterone Propionate ◽  
Female Rats ◽  
Male Rats ◽  
Male Rat ◽  
Peptidase Activity ◽  
Female Rat ◽  
Similar Relationship ◽  
Testosterone Treatment ◽  
Rat Hypothalamus

ABSTRACT It has previously been shown that the activity of certain peptidases in the female rat hypothalamus is related to the release of luteinizing hormone releasing factor from the tissue (Griffiths & Hooper 1972a). The activity of these enzymes was investigated after orchidectomy and testosterone propionate injection to determine if a similar relationship exists in male rats. The depression in supernatant activity following orchidectomy and the elevation after testosterone treatment are interpreted as confirming this, and it is proposed that alterations in peptidase activity may be used as an index of gonadotrophin release in male as well as in female rats.

CHANGES IN HYPOTHALAMIC PEPTIDASE ACTIVITY DURING THE OESTROUS CYCLE IN THE ADULT FEMALE RAT

1973 ◽  
Vol 74 (1) ◽  
pp. 41-48 ◽  
Author(s):  
E. C. Griffiths ◽  
K. C. Hooper
Keyword(s):  
Enzyme Activity ◽  
Luteinizing Hormone ◽  
Oestrous Cycle ◽  
Female Rats ◽  
Male Rats ◽  
Supernatant Fraction ◽  
Normal Male ◽  
Normal Female ◽  
Peptidase Activity ◽  
Female Rat

ABSTRACT The activity of peptidases in the rat hypothalamus which are capable of inactivating oxytocin has previously been found to vary with stimuli known to influence gonadotrophin release and may be related to both luteinizing hormone (LH) and luteinizing hormone releasing factor (LH-RF) release (Griffith & Hooper 1972a,b). In the present study, enzyme activity was determined in normal female rats during the morning and afternoon of each stage of the oestrous cycle, in normal rats, and in female rats injected neonatally with testosterone. The activity of the supernatant fraction was found to be not significantly different during the morning of each stage, but was greatly decreased on the afternoon of pro-oestrus; particulate activity did not vary during the oestrous cycle. Supernatant and particulate activities were found to be the same in normal male rats and testosterone-treated females, as previously shown. Both fractions' activities were significantly less than those found in the oestrous cycle, other than on the afternoon of pro-oestrus. These results indicate changes in hypothalamic peptidase activity during the oestrous cycle which may be inversely related to LH and LH-RF release; they also confirm the masculinizing effect of neonatal testosterone on the hypothalamus.

PEPTIDASE ACTIVITY IN DIFFERENT AREAS OF THE RAT HYPOTHALAMUS

1974 ◽  
Vol 77 (1) ◽  
pp. 10-18 ◽  
Author(s):  
E. C. Griffiths ◽  
K. C. Hooper
Keyword(s):  
Gonadal Steroids ◽  
Female Rats ◽  
Supernatant Fraction ◽  
Normal Male ◽  
Peptidase Activity ◽  
Gonadal Steroid ◽  
Rat Hypothalamus ◽  
Selective Distribution ◽  
Male And Female Rats ◽  
Hypothalamic Level

ABSTRACT To provide further information on the function of peptidases present in the rat hypothalamus which are capable of inactivating oxytocin, these enzymes' activity was investigated in anterior, middle and posterior hypothalamic areas of normal male and female rats, gonadectomized rats and rats gonadectomized and injected with gonadal steroids (oestradiol in female animals, testosterone in males). Peptidase activity in the supernatant fraction was distributed unevenly through the hypothalamus with the majority of activity in the anterior and middle areas from both sexes; particulate peptidase activity occurred principally in the middle and posterior areas. Gonadectomy selectively decreased supernatant activity in the anterior and middle areas, whereas steroid treatment reversed this effect, but neither caused any change in particulate activity from female rats and only small changes in male animals. These results are interpreted as indicating a selective distribution of supernatant peptidase activity in those hypothalamic areas responsible for luteinizing hormone – releasing factor (LRF) synthesis and release, and confirming previous findings that this fraction may be involved in LRF metabolism. They may also suggest the sites of gonadal steroid feedback at the hypothalamic level.

PEPTIDASE ACTIVITY IN THE HYPOTHALAMI OF RATS TREATED NEONATALLY WITH OESTROGEN

1973 ◽  
Vol 72 (1) ◽  
pp. 9-17 ◽  
Author(s):  
E. C. Griffiths ◽  
K. C. Hooper
Keyword(s):  
Sexual Differentiation ◽  
Critical Period ◽  
Testosterone Propionate ◽  
Marked Decrease ◽  
Reproductive Function ◽  
Female Rats ◽  
Male Rats ◽  
Peptidase Activity ◽  
Rat Hypothalamus ◽  
Lh Secretion

ABSTRACT The previous paper (Griffiths & Hooper 1973) described the activity of certain hypothalamic peptidases following orchidectomy and testosterone propionate injection, and suggested that changes in enzyme levels may be used as an index of gonadotrophin release in male rats, in a similar way to that previously described for female rats (Griffiths & Hooper 1972a). Using this approach, the effect of neonatally administered oestrogen on the hypothalamus was investigated. The marked increase in supernatant activity in male rats and the equally marked decrease in supernatant activity in female rats, both injected during the critical period of hypothalamic sexual differentiation, are interpreted as indicating decreased LH secretion in males and increased LH secretion in females respectively. It can be concluded that the changes in reproductive function produced by neonatally administered oestrogen are caused by alterations in LH-RF metabolism and that the peptidases in the rat hypothalamus are responsible for this metabolism.

scholarly journals Photoenzymatic Repair of Ultraviolet Damage in DNA

1962 ◽  
Vol 45 (4) ◽  
pp. 703-724 ◽  
Author(s):  
Claud S. Rupert
Keyword(s):  
Dna Repair ◽  
Competitive Inhibition ◽  
Kinetic Studies ◽  
Reaction Scheme ◽  
Substrate Complex ◽  
Enzyme Substrate ◽  
Light Intensities ◽  
Presence And Absence ◽  
Ultraviolet Damage ◽  
Kinetics Of

As previously reported, ultraviolet-inactivated bacterial transforming DNA can be restored to activity by an enzyme-like agent from bakers' yeast which requires light for its activity. Kinetics of this reaction, in the presence and absence of inhibitors, are found consistent with the Michaelis-Menten reaction scheme, with the sites of ultraviolet damage on the DNA serving as substrate and the repaired structure as product. Kinetic studies with different light intensities suggest that the necessary illumination causes photolysis of the enzyme-substrate complex with concurrent repair of the DNA. Competitive inhibition of irradiated transforming DNA repair, which occurs when irradiated non-transforming DNA is present in the same reaction mixture, permits ultraviolet damage (of the kind capable of being photoreactivated) to be detected in any type of DNA.

Kinetic Behaviour of Amylase According to pH: A New Perspective for Starch Hydrolysis Process

2020 ◽  
Vol 16 (2) ◽  
pp. 135-144
Author(s):  
Ravneet K. Grewal ◽  
Baldeep Kaur ◽  
Gagandeep Kaur
Keyword(s):  
Metal Ions ◽  
Competitive Inhibition ◽  
Deae Cellulose ◽  
Kinetic Studies ◽  
Cost Effective ◽  
Starch Hydrolysis ◽  
Divalent Metal Ions ◽  
Activity Data ◽  
Time Saving ◽  
Alkaline Conditions

Background: Amylases are the most widely used biocatalysts in starch saccharification and detergent industries. However, commercially available amylases have few limitations viz. limited activity at low or high pH and Ca2+ dependency. Objective: The quest for exploiting amylase for diverse applications to improve the industrial processes in terms of efficiency and feasibility led us to investigate the kinetics of amylase in the presence of metal ions as a function of pH. Methods: The crude extract from soil fungal isolate cultures is subjected to salt precipitation, dialysis and DEAE cellulose chromatography followed by amylase extraction and is incubated with divalent metal ions (i.e., Ca2+, Fe2+, Cu2+, and Hg2+); Michaelis-Menton constant (Km), and maximum reaction velocity (Vmax) are calculated by plotting the activity data obtained in the absence and presence of ions, as a function of substrate concentration in Lineweaver-Burk Plot. Results: Kinetic studies reveal that amylase is inhibited un-competitively at 5mM Cu2+ at pH 4.5 and 7.5, but non-competitively at pH 9.5. Non-competitive inhibition of amylase catalyzed starch hydrolysis is observed with 5mM Hg2+ at pH 9.5, which changes to mixed inhibition at pH 4.5 and 7.5. At pH 4.5, Ca2+ induces K- and V-type activation of amylase catalyzed starch hydrolysis; however, the enzyme has V-type activation at 7mM Ca2+ under alkaline conditions. Also, K- and V-type of activation of amylase is observed in the presence of 7mM Fe2+ at pH 4.5 and 9.5. Conclusion: These findings suggest that divalent ions modulation of amylase is pH dependent. Furthermore, a time-saving and cost-effective solution is proposed to overcome the challenges of the existing methodology of starch hydrolysis in starch and detergent industries.

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