TY - JOUR
T1 - Substrate-based inhibitors of the (S)-adenosyl-L-methionine:Δ(24(25))- to Δ(24(28))-sterol methyl transferase from Saccharomyces cerevisiae
AU - Nes, W. David
AU - Guo, De an
AU - Zhou, Wen
N1 - Funding Information:
We thank Ya-jun Cui, Darrel Morrison (supported by NSF-REU Fellowship BIR-9322342), and Zhonghua Jia for technical support. The organic chemistry study on the synthesis, puri®cation, and characterization of sterol methylation inhibitors serves to ful®ll part of the Ph.D. requirements of Wen Zhou.
Funding Information:
1 This research was supported by Welch Foundation Grant D-1276 (W.D.N.). 2To whom correspondence should be addressed.
PY - 1997/6/1
Y1 - 1997/6/1
N2 - A series of 31 side-chain-modified analogs of cholesterol, zymosterol, lanosterol, and cycloartenol and the steroidal alkaloids solasodine and solanidine were studied as inhibitors of (S)-adenosyl-L- methionine:Δ(24(25))-sterol methyl transferase (SMT) enzyme activity from Saccharomyces cerevisiae. Two classes of sterol methylation inhibitors were tested: substrate analogs, including mechanism-based inhibitors, and transition state analogs. Several novel sterol methylation inhibitors that contained an aza, aziridine, or ammonium group in the sterol side chain were prepared and tested for the first time. The degree and kinetic pattern of methylation inhibition were found to be influenced by the position and nature of the variant functional group introduced into the side chain. The most potent inhibitors of SMT enzyme activity were transition state analog inhibitors (K(i) values of 5 to 10 nM) that mimicked the structure and conformation of the natural substrate presumed to form in the ternary complex generated in the transition state. Steroidal alkaloids were potent competitive inhibitors with K(i) values ranging from 2 to 30 μM, which is about the K(mapp) of zymosteroi, ca. 27 μM. An isosteric analog of the natural substrate, zymosterol, in which the 26/27-gem-dimethyl groups were joined to form a cyclopropylidene function is shown to be a potent irreversible mechanism-based inactivator of SMT enzyme activity that exhibits competitive-type inhibition, K(i) 48 μM with a K(inact) of 1.52 min-1. Mechanistic implications of these results provide new insights into the topology of the ternary complex involving sterol-AdoMet-enzyme.
AB - A series of 31 side-chain-modified analogs of cholesterol, zymosterol, lanosterol, and cycloartenol and the steroidal alkaloids solasodine and solanidine were studied as inhibitors of (S)-adenosyl-L- methionine:Δ(24(25))-sterol methyl transferase (SMT) enzyme activity from Saccharomyces cerevisiae. Two classes of sterol methylation inhibitors were tested: substrate analogs, including mechanism-based inhibitors, and transition state analogs. Several novel sterol methylation inhibitors that contained an aza, aziridine, or ammonium group in the sterol side chain were prepared and tested for the first time. The degree and kinetic pattern of methylation inhibition were found to be influenced by the position and nature of the variant functional group introduced into the side chain. The most potent inhibitors of SMT enzyme activity were transition state analog inhibitors (K(i) values of 5 to 10 nM) that mimicked the structure and conformation of the natural substrate presumed to form in the ternary complex generated in the transition state. Steroidal alkaloids were potent competitive inhibitors with K(i) values ranging from 2 to 30 μM, which is about the K(mapp) of zymosteroi, ca. 27 μM. An isosteric analog of the natural substrate, zymosterol, in which the 26/27-gem-dimethyl groups were joined to form a cyclopropylidene function is shown to be a potent irreversible mechanism-based inactivator of SMT enzyme activity that exhibits competitive-type inhibition, K(i) 48 μM with a K(inact) of 1.52 min-1. Mechanistic implications of these results provide new insights into the topology of the ternary complex involving sterol-AdoMet-enzyme.
UR - http://www.scopus.com/inward/record.url?scp=0031172883&partnerID=8YFLogxK
U2 - 10.1006/abbi.1997.9984
DO - 10.1006/abbi.1997.9984
M3 - Article
C2 - 9185615
AN - SCOPUS:0031172883
VL - 342
SP - 68
EP - 81
JO - Archives of Biochemistry and Biophysics
JF - Archives of Biochemistry and Biophysics
SN - 0003-9861
IS - 1
ER -