Purified human leukotriene A4 hydrolase is shown to contain 1 mol of zinc per mol of enzyme, as determined by atomic absorption spectrometry. The enzyme is inhibited dose-dependently by the chelating agents 8-hydroxy-quinoline-5-sulfonic acid, and 1,10-phenanthroline with KI values of about 2 and 8 x 10(-4) M, respectively, whereas dipicolinic acid and EDTA are ineffective in this respect. The inhibition by 1,10-phenanthroline is time-dependent, and at a concentration of 5 mM, 50% inhibition of enzyme (3 x 10(-7) M) occurs after about 15 min. The zinc atom of leukotriene A4 hydrolase can be removed by dialysis against 1,10-phenanthroline which results in loss of enzyme activity. The catalytic activity is almost completely restored by the addition of stoichiometric amounts of Zn2+ or Co2+.

Amino Acid Sequence↗

PubMed1990

Mutagenesis of residues flanking Lys-40 enhances the enzymatic activity and reduces the angiogenic potency of angiogenin.

The primary structure of the blood vessel inducing protein angiogenin is 35% identical with that of pancreatic ribonuclease (RNase) and contains counterparts for the critical RNase active-site residues His-12, Lys-41, and His-119. Although angiogenin is a ribonucleolytic enzyme, its activity toward conventional substrates is lower than that of pancreatic RNase by several orders of magnitude. Comparison of the amino acid sequences of RNase and angiogenin reveals several striking differences in the region flanking the active-site lysine, including a deletion and a transposition of aspartic acid and proline residues. In order to examine how these sequence changes alter the functional properties of angiogenin, an angiogenin/RNase hybrid protein (ARH-II), in which residues 38-41 of angiogenin (Pro-Cys-Lys-Asp) have been replaced by the corresponding segment of bovine pancreatic RNase (Asp-Arg-Cys-Lys-Pro), was prepared by regional mutagenesis. Compared to angiogenin, ARH-II has markedly diminished angiogenic activity on the chick embryo chorioallantoic membrane but 5-75-fold greater enzymatic activity toward a variety of polynucleotide and dinucleotide substrates. In addition, the specificity of ARH-II toward dinucleotide substrates differs from that of angiogenin and is qualitatively similar to that of pancreatic RNase. Thus, non-active-site residues near Lys-40 in angiogenin appear to play a significant role in determining enzymatic specificity and reactivity as well as angiogenic potency. An additional angiogenin/RNase hybrid protein (ARH-IV), in which residues 59-71 of ARH-II have been replaced by the corresponding segment of pancreatic RNase, was also prepared.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1990

Kidney transplantation under FK 506.

The experimental immunosuppressive drug FK 506 was given to 36 renal transplant recipients, many of whom were highly sensitized. Ten were undergoing kidney retransplantation, 10 also underwent liver transplantation at an earlier time (6 patients) or concomitantly (4 patients), and 2 patients received a third organ (heart or pancreas) in addition to a liver and kidney. With follow-ups of 4 to 13 months, all but 2 of the 36 patients are alive, 29 (81%) are dialysis free, and most have good renal function. Twenty of the 29 dialysis-free patients are receiving no or low-dose (2.5 to 5.0 mg/d) prednisone therapy. Only one kidney was lost to cellular rejection. However, patients who had antidonor cytotoxic antibodies in current or historical serum samples had a high rate (3 of 9) of irreversible humoral rejection. A low incidence of posttransplant hypertension was noteworthy. Hirsutism and gingival hyperplasia were not observed. Serum cholesterol levels in patients who took FK 506 were unexpectedly low, and the effect on the level of uric acid was minimal. The side effects of FK 506 therapy include nephrotoxicity, neurotoxicity, and potential induction of a diabetic state. These are similar to the side effects of cyclosporine use, but probably less severe. The seeming safety, efficacy, and relative freedom from side effects of FK 506 encourage further trials in kidney transplantation.

Adult↗

PubMed1990

Liver, kidney, and thoracic organ transplantation under FK 506.

The new immunosuppressive drug FK 506 was used from the outset with low doses of prednisone to treat 120 recipients of primary liver grafts and 20 more patients undergoing liver retransplantation. The patient survival rate after 2 to 8 months in the primary liver transplantation series is 93.3%, with original graft survival of 87.5%. Of the 20 patients in the hepatic retransplant series, 17 (85%) are living. Almost all of the surviving patients have good liver function. In addition 11 hearts, 2 double lungs, and a heart-lung have been transplanted under FK 506, with survival of all 14 patients. With all of the organ systems so far tested, including the kidney (which has been reported elsewhere), rejection usually has been controlled without additional drugs and with lower average steroid doses than in the past. Nephrotoxicity has been observed, but not to an alarming degree, and there has been a notable absence of hypertension. There is a suggestion that serum cholesterol may be lowered by FK 506, but this is unproved. Although the adverse reactions of FK 506 and the immunosuppressive mechanisms resemble those of cyclosporine, our preliminary observations suggest that FK 506 may have a more advantageous therapeutic index.

Adolescent↗