Determination of azoferredoxin activity using cold-inactivated crude extracts of Clostridium pasteurianum

✍ Scribed by Esam Moustafa; L.E. Mortenson

• Book ID: 102625877
• Publisher: Elsevier Science
• Year: 1968
• Tongue: English
• Weight: 340 KB
• Volume: 24
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(68)90175-9

✦ Synopsis

Cell-free extracts from Clostridium pasteurianum catalyze the reduction of nitrogen and acetylene (l-4). Two protein components required for Nz fixation were isolated from these extracts. One component, molybdoferredoxin (MoFd) , contains molybdenum, nonhe'me iron and sulfide, and the other, azoferredoxin (AzoFd) , contains nonheme iron and sulfide (5). Since N,-fixing extracts from C. pa.steurianum when stored at temperatures near 0" lose their ability to catalyze nitrogen reduction (6), ATP-dependent hydrogen evolution in the presence of sodium dithionite (7), and acetylene reduction (as shown in this communication), we questioned whether either or both of these components was inactivated by storage at low temperatures. In the course of this work we have established that AzoFd is the component in these extracts that loses its activity when stored at O-1" (Fig. 1). Based on this finding and that acetylene reduction is valid as a method for measuring nitrogen fixation (24), a method was devised by which AzoFd activity was estimated by determining the rate of acetylene reduction catalyzed by cold-inactivated crude extracts to which AzoFd had been added. This paper describes the method of preparing the cold-inactivated extracts and the use of these extracts in determining the activity of AzoFd.

Methods

Extracts and Components of C. pasteurknum