Purification and characterization of Put1p from Saccharomyces cerevisiae.
In Saccharomyces cerevisiae, the PUT1 and PUT2 genes are required for the conversion of proline to glutamate. The PUT1 gene encodes Put1p, a proline dehydrogenase (PRODH) enzyme localized in the mitochondrion. Put1p was expressed and purified from Escherichia coli and shown to have a UV-visible absorption spectrum that is typical of a bound flavin cofactor. A K(m) value of 36 mM proline and a k(cat)=27 s(-1) were determined for Put1p using an artificial electron acceptor. Put1p also exhibited high activity using ubiquinone-1 (CoQ(1)) as an electron acceptor with a k(cat)=9.6 s(-1) and a K(m) of 33 microM for CoQ(1). In addition, knockout strains of the electron transfer flavoprotein (ETF) homolog in S. cerevisiae were able to grow on proline as the sole nitrogen source demonstrating that ETF is not required for proline utilization in yeast.
Wanduragala
Srimevan
S
Department of Biochemistry, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.
Sanyal
Nikhilesh
N
Liang
Xinwen
X
Becker
Donald F
DF
D004926:Escherichia coli / Q000235:genetics; D011392:Proline / Q000737:chemistry; D011394:Proline Oxidase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D014451:Ubiquinone / Q000737:chemistry
D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D014451:Ubiquinone; D011392:Proline; D011394:Proline Oxidase; C025203:Ubiquinone Q1
J Biol Chem. 1969 Aug 10;244(15):3999-4006
4389773
Mol Cell Biol. 1986 Jul;6(7):2638-45
3537723
Proc Natl Acad Sci U S A. 1988 Apr;85(7):2036-40
3353366
EMBO J. 2003 Oct 1;22(19):5147-53
14517252
J Bacteriol. 1979 Nov;140(2):504-7
387738
Nature. 1997 Sep 18;389(6648):300-5
9305847
J Nutr. 2008 Oct;138(10):2008S-2015S
18806116
Nat Struct Biol. 2003 Feb;10(2):109-14
12514740
J Proteome Res. 2006 Jul;5(7):1543-54
16823961
Anal Biochem. 1976 Aug;74(2):430-40
962101
J Biol Chem. 2005 Mar 25;280(12):11902-10
15665328
J Bacteriol. 1990 Sep;172(9):4927-35
1975578
Gene. 2002 May 15;290(1-2):1-18
12062797
J Biol Chem. 1993 Apr 25;268(12):8972-9
8473341
Appl Microbiol Biotechnol. 2008 Nov;81(2):211-23
18802692
Biochemistry. 2009 Feb 10;48(5):951-9
19140736
Arch Biochem Biophys. 2002 Dec 1;408(1):131-6
12485611
Amino Acids. 2008 Nov;35(4):719-30
18369526
Infect Immun. 2008 Jul;76(7):3037-44
18458068
Cancer Res. 2009 Aug 15;69(16):6414-22
19654292
J Bacteriol. 1979 Nov;140(2):498-503
387737
Biochemistry. 1979 Jan 23;18(2):331-7
570409
Biochemistry. 1995 Sep 12;34(36):11515-20
7547881
J Biol Chem. 2007 May 11;282(19):14316-27
17344208
J Clin Invest. 1992 Nov;90(5):1679-86
1430199
Mol Cell Biol. 1987 Dec;7(12):4431-40
3125423
EMBO J. 2003 Jul 15;22(14):3503-13
12853466
Oncogene. 2006 Sep 14;25(41):5640-7
16619034
Anal Biochem. 1989 Jul;180(1):136-9
2817336
Clin Chim Acta. 1976 Jan 16;66(2):227-39
1245071
J Biol Chem. 2009 Sep 4;284(36):24115-22
19574222
Biochemistry. 2005 Jun 28;44(25):9130-9
15966737
Cancer Res. 2001 Mar 1;61(5):1810-5
11280728
Annu Rev Biochem. 1980;49:1005-61
6250440
FEBS J. 2007 Nov;274(21):5481-504
17941859
Plant Physiol. 2004 Oct;136(2):3313-8
15448193
Biochemistry. 2004 Oct 5;43(39):12539-48
15449943
Biochemistry. 2001 Apr 17;40(15):4714-21
11294639
Biochemistry. 2008 Aug 26;47(34):8894-901
18672901