Complex of dipeptidyl peptidase II with adenosine deaminase.

Dipeptidyl peptidase II (DPPII) from bovine kidney cortex and lung was purified to the electrophoretically homogeneous state. The molecular and catalytic characteristics of the enzyme were determined. It was revealed that DPPII preparations possess adenosine deaminase (ADA) activity at all purification steps. For the first time, the ADA-binding ability of DPPII has been shown similar to the well-known ADA-binding enzyme, DPPIV. The dissociation constant of the DPPII-ADA complex was estimated using a resonant mirror biosensor (80 nM), fluorescence polarization (60 nM), and differential spectroscopy (36 nM) techniques. The data demonstrate that DPPII can form a complex with ADA, but with one order of magnitude higher dissociation constant than that of DPPIV (7.8 nM).

Interaction of adenosine deaminase with inhibitors. Chemical modification by diethyl pyrocarbonate.

The interaction of adenosine deaminase (adenosine aminohydrolase, ADA) from bovine spleen with inhibitors--erythro-9-(2-hydroxy-3-nonyl)adenine, erythro-9-(2-hydroxy-3-nonyl)-3-deazaadenine, and 1-deazaadenosine--was investigated. Using selective chemical modification by diethyl pyrocarbonate (DEP), the possible involvement of His residues in this interaction was studied. The graphical method of Tsou indicates that of six His residues modified in the presence of DEP, only one is essential for ADA activity. Inactivation of the enzyme, though with low rate, in complex with any of the inhibitors suggests that the adenine moiety of the inhibitors (and consequently, of the substrate) does not bind with the essential His to prevent its modification. The absence of noticeable changes in the dissociation constants of any of the enzyme-inhibitor complexes for the DEP-modified and control enzyme indicates that at least the most available His residues modified in our experiments do not participate in binding the inhibitors--derivatives of adenosine or erythro-9-(2-hydroxy-3-nonyl)adenine.

Activity of enzymes of adenyline compounds metabolism during crush and decompression of muscle tissue. Part I. Adenylate deaminase activity at experimental crush syndrome.

INTRODUCTION: The number of works on investigation of crush syndrome (CS) pathogenesis, of organs and enzymatic systems at traumatic toxicosis is rather limited. While the clinical current of trauma and the lethality prognosis depend on a degree of violations in them. Such investigations are necessary for opportune diagnosis and definition of a treatment tactic. To complete this deficiency, adenylate deaminase (AMPD) level was studied in twelve tissues at experimental CS in vivo. RESULTS: The experimental model of CS on white rats was caused by crush and decompression of femoral muscle tissue. The CS influence on AMPD activity was investigated in hemisphere, cerebellum, hypothalamus, pituitary body, heart, lung, liver, spleen, kidney, adrenal, crushed and not crushed muscles. In muscles, kidney, pituitary body and adrenal the activity decreased in 2 hours crush but the compensation of effect is observed after 5 hours crush. In cerebellum, hemisphere, heart, liver and lung it decreased during both of crush times. After 2 and 5 hours crush in hypothalamus and in spleen AMPD activity appeared much higher than in control. After 2 hours crush at the end of 48 hours decompression, the activity in muscles, kidney, adrenal and pituitary body was lower, in cerebellum, hypothalamus, hemisphere and heart--higher than, in lung, spleen and liver--close to control. After 5 hours crush in the majority of studied tissues at the end of decompression, the activity was below of control. The greatest deviance was observed in muscles. As to brain in cerebellum and hemisphere the parameter was close to, in pituitary body and in hypothalamus it was 1,5-fold of control. CONCLUSION: AMPD level in the most of studied tissues differs from the control at definite time of crush and decompression. These results mean the possibility of nucleotides pool balance distortion and intermediates accumulation.

Activity of enzymes of adenyline compounds metabolism during crush and decompression of muscle tissue. Part II. Adenosine deaminase activity at experimental crush syndrome.

INTRODUCTION: Publications on investigation of crush syndrome pathogenesis, particularly of enzymatic systems upon traumatic toxicosis are rather limited. Such investigations are necessary for opportune diagnosis and definition of a treatment tactic. To replenish this deficiency, the adenosine deaminase level was studied in 12 rat tissues at experimental crush syndrome in vivo. RESULTS: The experimental model of crush syndrome on white rats was induced by crush and decompression of femoral muscle tissue. The crush syndrome influence on activity of adenosine deaminase isoenzymes was investigated in hemisphere, cerebellum, hypothalamus, pituitary body, heart, lung, liver, spleen, kidney, adrenal, as well as in crushed and native muscles. In 2 and 5 hours after compression, the enzyme activity decreased in muscles, lung and heart; increased in hypothalamus; remains near the control value in kidney and spleen. In cerebellum the parameter practically does not vary during 2 hours compression, while increased in 5 hours. In adrenal, liver, pituitary body and hemisphere the data after 5 hours compression approximated the level of control value in account of compensating mechanism. In 48 hours decompression after 2 hours crush, the adenosine deaminase activity becomes higher than control value in hemisphere, hypothalamus, cerebellum, liver, heart, adrenal, intact muscle, lung and kidney; in the crushed muscle and spleen the activity is reduced down to 60% of control value. In 48 hours decompression after 5 hours compression, the enzyme activity is higher than control value in hypothalamus, pituitary body, hemisphere, cerebellum, kidney, adrenal, heart and lung. The activity is reduced in muscles, spleen and liver. CONCLUSION: The level of adenosine deaminase in most of studied tissues differs from the control value depending on compression and decompression time. It is worthy of note that namely during decompression, the enzyme level deviates from the control in the majority of tissues.

The effect of detergents on bovine adrenal ferredoxin.

The incubation of adrenal ferredoxin with various detergents in the presence of oxygen or ferricyanide leads to bleaching of the protein. The bleached preparation has the properties of apoferredoxin and it can be reconstituted with high yield by conventional methods.