Cys-92, Cys-95, and the C-terminal 12 residues of the Vibrio harveyi ferric uptake regulator (Fur) are functionally inessential.
Ferric uptake regulator (Fur) is a global regulator involved in multiple aspects of bacterial life. The gene encoding the Vibrio harveyi Fur (FurVh) was cloned from a pathogenic V. harveyi strain isolated from diseased fish. FurVh shares 77% overall sequence identity with the Escherichia coli Fur (FurEc) and could complement a mutant of FurEc. Like FurEc, FurVh, possesses two cysteine residues at positions 92 and 95, yet unlike FurEc, in which these cysteine residues constitute part of the metal ion coordination site and hence are vital to the repressor activity, C92 and C95 of FurVh proved to be functionally inessential. Further study identified a Vibrio Fur signature sequence, which is preserved in all the ten Vibrio Fur proteins that have been discovered to date but in none of the non-vibrio Fur proteins. Site-directed and random mutation analyses of the signature residues, the cysteine residues, and seven highly charged amino acid residues indicated that D9, H32, C137, and K138 of FurVh are functionally important but D9, C137, and K138 can be replaced by more than one functional substitutes. Systematic deletion analysis demonstrated that the C-terminal 12 residues of FurVh are functionally inessential. These results (i) indicated that the activation mechanism, or certain aspects of which, of FurVh is possibly different from that of FurEc; and (ii) suggested that it is not very likely that the C-terminal 12 residues play any significant role in the activation or stability of FurVh; and (iii) provided insights into the potential function of the local structure involving C137 and K138.
Sun
Kun
K
Institute of Oceanology, Chinese Academy of Sciences, Qingdao, 266071, P. R. China.
Cheng
Shuang
S
Zhang
Min
M
Wang
Fang
F
Sun
Li
L
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002681:China; D003545:Cysteine / Q000737:chemistry*; D005393:Fish Diseases / Q000382:microbiology*; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011817:Rabbits; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013329:Structure-Activity Relationship; D014733:Vibrio / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D014735:Vibrio Infections / Q000382:microbiology / Q000662:veterinary*
D001426:Bacterial Proteins; D012097:Repressor Proteins; C068477:ferric uptake regulating proteins, bacterial; D003545:Cysteine