产黄青霉的ATP硫酸化酶：酶的内部水平是否足以解释硫酸盐的利用速率？

ATP sulfurylase from Penicillium chrysogenum: is the internal level of the enzyme sufficient to account for the rate of sulfate utilization?

作者信息

Farley J R, Mayer S, Chandler C J, Segel I H

出版信息

J Bacteriol. 1979 Jan;137(1):350-6. doi: 10.1128/jb.137.1.350-356.1979.

DOI:10.1128/jb.137.1.350-356.1979

PMID:104967

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC218457/

Abstract

The in vivo rate of sulfate activation in Penicillium chrysogenum (wild-type strain ATCC 24791) was determined to be 0.19 +/- 0.09 mumol g(-1) (dry weight) min(-1) by the following methods. (i) The maximum growth of the organism in synthetic medium was a linear function of the initial Na(2)SO(4) concentration between 0 and 8 x 10(-4) Na(2)SO(4). The growth yield was 1.64 x 10(-2) g (dry weight) of mycelium per mumol of added sulfate, corresponding to a minimum sulfur requirement of 61 mumol/g (dry weight). Under these conditions (limiting sulfate) the minimum doubling time of P. chrysogenum in submerged culture was about 3.8 h, corresponding to a maximum exponential growth rate constant of 3.0 x 10(-3) min(-1). If all the sulfur in this mycelium passed through adenosine-5'-phosphosulfate, the rate of sulfate activation in vivo must have been 0.183 mumol min(-1) g(-1) (dry weight). (ii) In the presence of excess (35)SO(4) (2-), the total organic (35)S produced varied with the mycelial growth rate. However, until the culture approached maximum density, the product of [(growth rate constant) x (organic (35)S content)] was nearly constant at 0.24 to 0.28 mumol min(-1) g(-1) (dry weight). (iii) A sulfur-starved mycelium pulsed with 10(-4) M (35)SO(4) (2-) produced organic (35)S at a rate of about 0.10 mumol min(-1) g(-1) (dry weight) under conditions where the internal concentrations of ATP and sulfate would permit ATP sulfurylase to operate at about 70% of its V(max). Cell-free extracts of P. chrysogenum growing rapidly on excess sulfate contained 0.22 U of ATP sulfurylase per g (dry weight). Thus, in spite of the relatively low specific activity of homogeneous ATP sulfurylase (0.13 U/mg of protein, corresponding to an active site turnover of 7.15 min(-1)), the mycelial content of the enzyme was sufficient to account for the observed growth rate of the organism on inorganic sulfate as the sole sulfur source.

摘要

通过以下方法测定了产黄青霉（野生型菌株ATCC 24791）体内硫酸盐活化速率为0.19±0.09 μmol g⁻¹（干重）min⁻¹。（i）该生物体在合成培养基中的最大生长是0至8×10⁻⁴ Na₂SO₄之间初始Na₂SO₄浓度的线性函数。生长产量为每微摩尔添加的硫酸盐产生1.64×10⁻² g（干重）的菌丝体，对应最低硫需求量为61 μmol/g（干重）。在这些条件下（硫酸盐受限），产黄青霉在深层培养中的最短倍增时间约为3.8小时，对应最大指数生长速率常数为3.0×10⁻³ min⁻¹。如果该菌丝体中的所有硫都通过腺苷 - 5'-磷酸硫酸，那么体内硫酸盐活化速率必定为0.183 μmol min⁻¹ g⁻¹（干重）。（ii）在存在过量的³⁵SO₄²⁻时，产生的总有机³⁵S随菌丝体生长速率而变化。然而，直到培养物接近最大密度时，[（生长速率常数）×（有机³⁵S含量）]的乘积在0.24至0.28 μmol min⁻¹ g⁻¹（干重）之间几乎保持恒定。（iii）在内部ATP和硫酸盐浓度允许ATP硫酸化酶以其V(max)的约70%运行的条件下，用10⁻⁴ M ³⁵SO₄²⁻脉冲处理的缺硫菌丝体产生有机³⁵S的速率约为0.10 μmol min⁻¹ g⁻¹（干重）。在过量硫酸盐上快速生长的产黄青霉的无细胞提取物每克（干重）含有0.22 U的ATP硫酸化酶。因此，尽管均一的ATP硫酸化酶的比活性相对较低（0.13 U/mg蛋白质，对应活性位点周转数为7.15 min⁻¹），但该酶的菌丝体含量足以解释观察到的生物体以无机硫酸盐作为唯一硫源时的生长速率。